Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 25
Filtrar
Mais filtros

Base de dados
Tipo de documento
Intervalo de ano de publicação
1.
Lancet Oncol ; 21(5): 655-663, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32251621

RESUMO

BACKGROUND: We did a phase 2 trial of pembrolizumab in patients with non-small-cell lung cancer (NSCLC) or melanoma with untreated brain metastases to determine the activity of PD-1 blockade in the CNS. Interim results were previously published, and we now report an updated analysis of the full NSCLC cohort. METHODS: This was an open-label, phase 2 study of patients from the Yale Cancer Center (CT, USA). Eligible patients were at least 18 years of age with stage IV NSCLC with at least one brain metastasis 5-20 mm in size, not previously treated or progressing after previous radiotherapy, no neurological symptoms or corticosteroid requirement, and Eastern Cooperative Oncology Group performance status less than two. Modified Response Evaluation Criteria in Solid Tumors (mRECIST) criteria was used to evaluate CNS disease; systemic disease was not required for participation. Patients were treated with pembrolizumab 10 mg/kg intravenously every 2 weeks. Patients were in two cohorts: cohort 1 was for those with PD-L1 expression of at least 1% and cohort 2 was patients with PD-L1 less than 1% or unevaluable. The primary endpoint was the proportion of patients achieving a brain metastasis response (partial response or complete response, according to mRECIST). All treated patients were analysed for response and safety endpoints. This study is closed to accrual and is registered with ClinicalTrials.gov, NCT02085070. FINDINGS: Between March 31, 2014, and May 21, 2018, 42 patients were treated. Median follow-up was 8·3 months (IQR 4·5-26·2). 11 (29·7% [95% CI 15·9-47·0]) of 37 patients in cohort 1 had a brain metastasis response. There were no responses in cohort 2. Grade 3-4 adverse events related to treatment included two patients with pneumonitis, and one each with constitutional symptoms, colitis, adrenal insufficiency, hyperglycaemia, and hypokalaemia. Treatment-related serious adverse events occurred in six (14%) of 42 patients and were pneumonitis (n=2), acute kidney injury, colitis, hypokalaemia, and adrenal insufficiency (n=1 each). There were no treatment-related deaths. INTERPRETATION: Pembrolizumab has activity in brain metastases from NSCLC with PD-L1 expression at least 1% and is safe in selected patients with untreated brain metastases. Further investigation of immunotherapy in patients with CNS disease from NSCLC is warranted. FUNDING: Merck and the Yale Cancer Center.


Assuntos
Anticorpos Monoclonais Humanizados/administração & dosagem , Antígeno B7-H1/genética , Neoplasias Encefálicas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Idoso , Anticorpos Monoclonais Humanizados/efeitos adversos , Antígeno B7-H1/antagonistas & inibidores , Biomarcadores Tumorais/genética , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/secundário , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica
2.
Genet Med ; 19(7): 787-795, 2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28125075

RESUMO

PURPOSE: Implementing cancer precision medicine in the clinic requires assessing the therapeutic relevance of genomic alterations. A main challenge is the systematic interpretation of whole-exome sequencing (WES) data for clinical care. METHODS: One hundred sixty-five adults with metastatic colorectal and lung adenocarcinomas were prospectively enrolled in the CanSeq study. WES was performed on DNA extracted from formalin-fixed paraffin-embedded tumor biopsy samples and matched blood samples. Somatic and germ-line alterations were ranked according to therapeutic or clinical relevance. Results were interpreted using an integrated somatic and germ-line framework and returned in accordance with patient preferences. RESULTS: At the time of this analysis, WES had been performed and results returned to the clinical team for 165 participants. Of 768 curated somatic alterations, only 31% were associated with clinical evidence and 69% with preclinical or inferential evidence. Of 806 curated germ-line variants, 5% were clinically relevant and 56% were classified as variants of unknown significance. The variant review and decision-making processes were effective when the process was changed from that of a Molecular Tumor Board to a protocol-based approach. CONCLUSION: The development of novel interpretive and decision-support tools that draw from scientific and clinical evidence will be crucial for the success of cancer precision medicine in WES studies.Genet Med advance online publication 26 January 2017.


Assuntos
Sequenciamento do Exoma/métodos , Exoma/genética , Medicina de Precisão/métodos , Adenocarcinoma/genética , Adenocarcinoma de Pulmão , Adulto , Neoplasias Colorretais/genética , Bases de Dados Genéticas , Genômica/métodos , Mutação em Linhagem Germinativa/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Neoplasias Pulmonares/genética , Mutação/genética , Estudos Prospectivos , Análise de Sequência de DNA/métodos
3.
Proc Natl Acad Sci U S A ; 111(52): 18661-6, 2014 Dec 30.
Artigo em Inglês | MEDLINE | ID: mdl-25512530

RESUMO

Lung adenocarcinomas harboring activating mutations in the epidermal growth factor receptor (EGFR) represent a common molecular subset of non-small cell lung cancer (NSCLC) cases. EGFR mutations predict sensitivity to EGFR tyrosine kinase inhibitors (TKIs) and thus represent a dependency in NSCLCs harboring these alterations, but the genetic basis of EGFR dependence is not fully understood. Here, we applied an unbiased, ORF-based screen to identify genetic modifiers of EGFR dependence in EGFR-mutant NSCLC cells. This approach identified 18 kinase and kinase-related genes whose overexpression can substitute for EGFR in EGFR-dependent PC9 cells, and these genes include seven of nine Src family kinase genes, FGFR1, FGFR2, ITK, NTRK1, NTRK2, MOS, MST1R, and RAF1. A subset of these genes can complement loss of EGFR activity across multiple EGFR-dependent models. Unbiased gene-expression profiling of cells overexpressing EGFR bypass genes, together with targeted validation studies, reveals EGFR-independent activation of the MEK-ERK and phosphoinositide 3-kinase (PI3K)-AKT pathways. Combined inhibition of PI3K-mTOR and MEK restores EGFR dependence in cells expressing each of the 18 EGFR bypass genes. Together, these data uncover a broad spectrum of kinases capable of overcoming dependence on EGFR and underscore their convergence on the PI3K-AKT and MEK-ERK signaling axes in sustaining EGFR-independent survival.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/enzimologia , Receptores ErbB/biossíntese , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/enzimologia , Sistema de Sinalização das MAP Quinases , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Receptores ErbB/genética , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Glicoproteínas de Membrana/biossíntese , Glicoproteínas de Membrana/genética , Proteínas Tirosina Quinases/biossíntese , Proteínas Tirosina Quinases/genética , Proteínas Proto-Oncogênicas c-mos/biossíntese , Proteínas Proto-Oncogênicas c-mos/genética , Proteínas Proto-Oncogênicas c-raf/biossíntese , Proteínas Proto-Oncogênicas c-raf/genética , Receptores Proteína Tirosina Quinases/biossíntese , Receptores Proteína Tirosina Quinases/genética , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/biossíntese , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/biossíntese , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/genética , Receptor trkA/biossíntese , Receptor trkA/genética , Receptor trkB
5.
NPJ Precis Oncol ; 8(1): 3, 2024 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-38182677

RESUMO

EGFR tyrosine kinase inhibitors (TKIs) have transformed the treatment of EGFR-mutated non-small cell lung carcinoma (NSCLC); however, therapeutic resistance remains a clinical challenge. Acquired secondary EGFR mutations that increase ATP affinity and/or impair inhibitor binding are well-described mediators of resistance. Here we identify a de novo EGFR Y891D secondary alteration in a NSCLC with EGFR L858R. Acquired EGFR Y891D alterations were previously reported in association with resistance to first generation EGFR TKIs. Functional studies in Ba/F3 cells demonstrate reduced TKI sensitivity of EGFR L858R + Y891D, with the greatest reduction observed for first and second generation TKIs. Unlike other EGFR mutations associated with TKI resistance, Y891D does not significantly alter ATP affinity or promote steric hindrance to inhibitor binding. Our data suggest that the Y891D mutation destabilizes EGFR L858R, potentially generating a population of misfolded receptor with preserved signaling capacity but reduced sensitivity to EGFR inhibitors. These findings raise the possibility of protein misfolding as a mechanism of resistance to EGFR inhibition in EGFR-mutated NSCLC.

6.
Blood ; 117(9): 2567-76, 2011 Mar 03.
Artigo em Inglês | MEDLINE | ID: mdl-21068437

RESUMO

Haploinsufficiency for ribosomal protein genes has been implicated in the pathophysiology of Diamond-Blackfan anemia (DBA) and the 5q-syndrome, a subtype of myelodysplastic syndrome. The p53 pathway is activated by ribosome dysfunction, but the molecular basis for selective impairment of the erythroid lineage in disorders of ribosome function has not been determined. We found that p53 accumulates selectively in the erythroid lineage in primary human hematopoietic progenitor cells after expression of shRNAs targeting RPS14, the ribosomal protein gene deleted in the 5q-syndrome, or RPS19, the most commonly mutated gene in DBA. Induction of p53 led to lineage-specific accumulation of p21 and consequent cell cycle arrest in erythroid progenitor cells. Pharmacologic inhibition of p53 rescued the erythroid defect, whereas nutlin-3, a compound that activates p53 through inhibition of HDM2, selectively impaired erythropoiesis. In bone marrow biopsies from patients with DBA or del(5q) myelodysplastic syndrome, we found an accumulation of nuclear p53 staining in erythroid progenitor cells that was not present in control samples. Our findings indicate that the erythroid lineage has a low threshold for the induction of p53, providing a basis for the failure of erythropoiesis in the 5q-syndrome, DBA, and perhaps other bone marrow failure syndromes.


Assuntos
Células Precursoras Eritroides/metabolismo , Haploinsuficiência/genética , Proteínas Ribossômicas/genética , Proteína Supressora de Tumor p53/metabolismo , Anemia de Diamond-Blackfan/genética , Anemia de Diamond-Blackfan/patologia , Anemia Macrocítica/genética , Anemia Macrocítica/patologia , Animais , Benzotiazóis/farmacologia , Ciclo Celular/efeitos dos fármacos , Linhagem da Célula/efeitos dos fármacos , Nucléolo Celular/efeitos dos fármacos , Nucléolo Celular/metabolismo , Deleção Cromossômica , Cromossomos Humanos Par 5/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Células Precursoras Eritroides/efeitos dos fármacos , Células Precursoras Eritroides/patologia , Hematopoese/efeitos dos fármacos , Humanos , Imidazóis/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Síndromes Mielodisplásicas/genética , Síndromes Mielodisplásicas/patologia , Piperazinas/metabolismo , Ligação Proteica/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , RNA Interferente Pequeno/metabolismo , Proteínas Ribossômicas/deficiência , Proteínas Ribossômicas/metabolismo , Tolueno/análogos & derivados , Tolueno/farmacologia
7.
Nat Genet ; 35(4): 372-6, 2003 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-14608358

RESUMO

A key question in systems biology is how diverse physiologic processes are integrated to produce global homeostasis. Genetic analysis can contribute by identifying genes that perturb this integration. One system orchestrates renal NaCl and K+ flux to achieve homeostasis of blood pressure and serum K+ concentration. Positional cloning implicated the serine-threonine kinase WNK4 in this process; clustered mutations in PRKWNK4, encoding WNK4, cause hypertension and hyperkalemia (pseudohypoaldosteronism type II, PHAII) by altering renal NaCl and K+ handling. Wild-type WNK4 inhibits the renal Na-Cl cotransporter (NCCT); mutations that cause PHAII relieve this inhibition. This explains the hypertension of PHAII but does not account for the hyperkalemia. By expression in Xenopus laevis oocytes, we show that WNK4 also inhibits the renal K+ channel ROMK. This inhibition is independent of WNK4 kinase activity and is mediated by clathrin-dependent endocytosis of ROMK, mechanisms distinct from those that characterize WNK4 inhibition of NCCT. Most notably, the same mutations in PRKWNK4 that relieve NCCT inhibition markedly increase inhibition of ROMK. These findings establish WNK4 as a multifunctional regulator of diverse ion transporters; moreover, they explain the pathophysiology of PHAII. They also identify WNK4 as a molecular switch that can vary the balance between NaCl reabsorption and K+ secretion to maintain integrated homeostasis.


Assuntos
Rim/fisiologia , Canais de Potássio Corretores do Fluxo de Internalização , Potássio/metabolismo , Proteínas Serina-Treonina Quinases/fisiologia , Receptores de Droga , Cloreto de Sódio/metabolismo , Simportadores , Animais , Proteínas de Transporte/antagonistas & inibidores , Proteínas de Transporte/metabolismo , Clatrina/metabolismo , Endocitose , Proteínas de Fluorescência Verde , Transporte de Íons , Proteínas Luminescentes/metabolismo , Camundongos , Canais de Potássio/metabolismo , Pseudo-Hipoaldosteronismo/metabolismo , Ratos , Simportadores de Cloreto de Sódio , Membro 3 da Família 12 de Carreador de Soluto , Xenopus laevis/metabolismo
8.
Oncology ; 82(3): 165-7, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22433430

RESUMO

Chemotherapy-induced peripheral neurotoxicity is a major problem because it represents the dose-limiting side effect of a significant number of antineoplastic drugs, such as vinca alkaloids. Hereditary neuropathies usually predispose to severe vincristine neurotoxicity. Here, we report the case of a 56-year-old man with Machado-Joseph disease, also known as spinocerebellar ataxia type 3, treated with a vinca alkaloid without exacerbation of neurological symptoms.


Assuntos
Antineoplásicos Fitogênicos/administração & dosagem , Doença de Machado-Joseph/tratamento farmacológico , Vincristina/administração & dosagem , Humanos , Masculino , Pessoa de Meia-Idade , Imagem Multimodal , Tomografia por Emissão de Pósitrons , Prognóstico , Tomografia Computadorizada por Raios X
9.
Clin Cancer Res ; 28(14): 3091-3103, 2022 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-35247929

RESUMO

PURPOSE: The identification of actionable oncogenic alterations has enabled targeted therapeutic strategies for subsets of patients with advanced malignancies, including lung adenocarcinoma (LUAD). We sought to assess the frequency of known drivers and identify new candidate drivers in a cohort of LUAD from patients with minimal smoking history. EXPERIMENTAL DESIGN: We performed genomic characterization of 103 LUADs from patients with ≤10 pack-year smoking history. Tumors were subjected to targeted molecular profiling and/or whole-exome sequencing and RNA sequencing in search of established and previously uncharacterized candidate drivers. RESULTS: We identified an established oncogenic driver in 98 of 103 tumors (95%). From one tumor lacking a known driver, we identified a novel gene rearrangement between OCLN and RASGRF1. The encoded OCLN-RASGRF1 chimera fuses the membrane-spanning portion of the tight junction protein occludin with the catalytic RAS-GEF domain of the RAS activator RASGRF1. We identified a similar SLC4A4-RASGRF1 fusion in a pancreatic ductal adenocarcinoma cell line lacking an activating KRAS mutation and an IQGAP1-RASGRF1 fusion from a sarcoma in The Cancer Genome Atlas. We demonstrate these fusions increase cellular levels of active GTP-RAS, induce cellular transformation, and promote in vivo tumorigenesis. Cells driven by RASGRF1 fusions are sensitive to targeting of the RAF-MEK-ERK pathway in vitro and in vivo. CONCLUSIONS: Our findings credential RASGRF1 fusions as a therapeutic target in multiple malignancies and implicate RAF-MEK-ERK inhibition as a potential treatment strategy for advanced tumors harboring these alterations. See related commentary by Moorthi and Berger, p. 2983.


Assuntos
Adenocarcinoma de Pulmão , Neoplasias Pulmonares , Carcinogênese/genética , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Quinases de Proteína Quinase Ativadas por Mitógeno , ras-GRF1/genética
10.
JMIR Cancer ; 8(2): e33859, 2022 05 19.
Artigo em Inglês | MEDLINE | ID: mdl-35588361

RESUMO

BACKGROUND: Expert knowledge is often shared among multidisciplinary academic teams at tumor boards (TBs) across the country, but these conversations exist in silos and do not reach the wider oncology community. OBJECTIVE: Using an oncologist-only question and answer (Q&A) website, we sought to document expert insights from TBs at National Cancer Institute-designated Comprehensive Cancer Centers (NCI-CCCs) to provide educational benefits to the oncology community. METHODS: We designed a process with the NCI-CCCs to document and share discussions from the TBs focused on areas of practice variation on theMednet, an interactive Q&A website of over 13,000 US oncologists. The faculty translated the TB discussions into concise, non-case-based Q&As on theMednet. Answers were peer reviewed and disseminated in email newsletters to registered oncologists. Reach and engagement were measured. Following each Q&A, a survey question asked how the TB Q&As impacted the readers' practice. RESULTS: A total of 23 breast, thoracic, gastrointestinal, and genitourinary programs from 16 NCI-CCC sites participated. Between December 2016 and July 2021, the faculty highlighted 368 questions from their TBs. Q&As were viewed 147,661 times by 7381 oncologists at 3515 institutions from all 50 states. A total of 277 (75%) Q&As were viewed every month. Of the 1063 responses to a survey question on how the Q&A affected clinicians' practices, 646 (61%) reported that it confirmed their current practice, 163 (20%) indicated that a Q&A would change their future practice, and 214 (15%) reported learning something new. CONCLUSIONS: Through an online Q&A platform, academics at the NCI-CCCs share knowledge outside the walls of academia with oncologists across the United States. Access to up-to-date expert knowledge can reassure clinicians' practices, significantly impact patient care in community practices, and be a source of new knowledge and education.

11.
Cancer Discov ; 8(6): 676-678, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29858224

RESUMO

A patient with advanced lung adenocarcinoma harboring a CD74-NRG1 gene rearrangement, which promotes ERBB2-ERBB3 heterodimerization and activation of downstream signaling, had an exceptional therapeutic response to an experimental anti-ERBB3 antibody. This result illustrates how NRG1 rearrangements, which are observed at a low frequency in a variety of solid tumors, may represent tractable therapeutic targets. Cancer Discov; 8(6); 676-8. ©2018 AACR.See related article by Drilon et al., p. 686.


Assuntos
Adenocarcinoma de Pulmão , Neuregulina-1 , Linhagem Celular Tumoral , Humanos , Ligantes , Receptor ErbB-2 , Receptor ErbB-3 , Transdução de Sinais
12.
Clin Cancer Res ; 24(23): 5963-5976, 2018 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-30072474

RESUMO

PURPOSE: MET inhibitors can be effective therapies in patients with MET exon 14 (METex14) mutant non-small cell lung cancer (NSCLC). However, long-term efficacy is limited by the development of drug resistance. In this study, we characterize acquired amplification of wild-type (WT) KRAS as a molecular mechanism behind crizotinib resistance in three cases of METex14-mutant NSCLC and propose a combination therapy to target it. EXPERIMENTAL DESIGN: The patient-derived cell line and xenograft (PDX) DFCI358 were established from a crizotinib-resistant METex14-mutant patient tumor with massive focal amplification of WT KRAS. To characterize the mechanism of KRAS-mediated resistance, molecular signaling was analyzed in the parental cell line and its KRAS siRNA-transfected derivative. Sensitivity of the cell line to ligand stimulation was assessed and KRAS-dependent expression of EGFR ligands was quantified. Drug combinations were screened for efficacy in vivo and in vitro using viability and apoptotic assays. RESULTS: KRAS amplification is a recurrent genetic event in crizotinib-resistant METex14-mutant NSCLC. The key characteristics of this genetic signature include uncoupling MET from downstream effectors, relative insensitivity to dual MET/MEK inhibition due to compensatory induction of PI3K signaling, KRAS-induced expression of EGFR ligands and hypersensitivity to ligand-dependent and independent activation, and reliance on PI3K signaling upon MET inhibition. CONCLUSIONS: Using patient-derived cell line and xenografts, we characterize the mechanism of crizotinib resistance mediated by KRAS amplification in METex14-mutant NSCLC and demonstrate the superior efficacy of the dual MET/PI3K inhibition as a therapeutic strategy addressing this resistance mechanism.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Resistencia a Medicamentos Antineoplásicos/genética , Éxons , Amplificação de Genes , Neoplasias Pulmonares/genética , Mutação , Proteínas Proto-Oncogênicas c-met/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Animais , Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Linhagem Celular Tumoral , Crizotinibe/farmacologia , Variações do Número de Cópias de DNA , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica , Humanos , Hibridização in Situ Fluorescente , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/tratamento farmacológico , Camundongos , Modelos Biológicos , Fosfatidilinositol 3-Quinases/genética , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , Inibidores de Proteínas Quinases/farmacologia , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
13.
Trends Endocrinol Metab ; 16(3): 98-103, 2005 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-15808806

RESUMO

Key components of complex physiological regulatory pathways can be uncovered through the molecular-genetic study of rare, inherited diseases. WNK kinases are a recently discovered class of serine-threonine kinases that are distinctive because of the substitution of cysteine for lysine in subdomain II of the catalytic domain. Mutations in PRKWNK1 and PRKWNK4, which encode WNK1 and WNK4, result in an inherited syndrome of hypertension and hyperkalemia. Recent physiological work has revealed that WNK4 alters the balance of NaCl reabsorption and K(+) secretion in the distal nephron by actions on both transcellular and paracellular ion-flux pathways. Additionally, WNK4 is expressed in extra-renal epithelia with prominent roles in Cl(-) handling, and it regulates transporters that are responsible for Cl(-) flux across apical and basolateral membranes. WNK kinases are components of a novel signaling pathway that is important for the control of blood pressure and electrolyte homeostasis.


Assuntos
Íons/metabolismo , Proteínas Serina-Treonina Quinases/fisiologia , Animais , Bioquímica/métodos , Humanos , Transporte de Íons/fisiologia , Biologia Molecular/métodos
14.
Science ; 351(6278): 1214-8, 2016 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-26912360

RESUMO

The discovery of cancer dependencies has the potential to inform therapeutic strategies and to identify putative drug targets. Integrating data from comprehensive genomic profiling of cancer cell lines and from functional characterization of cancer cell dependencies, we discovered that loss of the enzyme methylthioadenosine phosphorylase (MTAP) confers a selective dependence on protein arginine methyltransferase 5 (PRMT5) and its binding partner WDR77. MTAP is frequently lost due to its proximity to the commonly deleted tumor suppressor gene, CDKN2A. We observed increased intracellular concentrations of methylthioadenosine (MTA, the metabolite cleaved by MTAP) in cells harboring MTAP deletions. Furthermore, MTA specifically inhibited PRMT5 enzymatic activity. Administration of either MTA or a small-molecule PRMT5 inhibitor showed a modest preferential impairment of cell viability for MTAP-null cancer cell lines compared with isogenic MTAP-expressing counterparts. Together, our findings reveal PRMT5 as a potential vulnerability across multiple cancer lineages augmented by a common "passenger" genomic alteration.


Assuntos
Neoplasias/tratamento farmacológico , Proteína-Arginina N-Metiltransferases/metabolismo , Purina-Núcleosídeo Fosforilase/metabolismo , Linhagem Celular Tumoral , Desoxiadenosinas/metabolismo , Desoxiadenosinas/farmacologia , Inibidores Enzimáticos/farmacologia , Deleção de Genes , Humanos , Isoquinolinas/farmacologia , Neoplasias/enzimologia , Proteína-Arginina N-Metiltransferases/antagonistas & inibidores , Proteína-Arginina N-Metiltransferases/genética , Purina-Núcleosídeo Fosforilase/genética , Pirimidinas/farmacologia , Tionucleosídeos/metabolismo , Tionucleosídeos/farmacologia , Fatores de Transcrição
15.
Cancer Cell ; 27(3): 397-408, 2015 Mar 09.
Artigo em Inglês | MEDLINE | ID: mdl-25759024

RESUMO

We conducted a large-scale functional genetic study to characterize mechanisms of resistance to ALK inhibition in ALK-dependent lung cancer cells. We identify members of known resistance pathways and additional putative resistance drivers. Among the latter were members of the P2Y purinergic receptor family of G-protein-coupled receptors (P2Y1, P2Y2, and P2Y6). P2Y receptors mediated resistance in part through a protein-kinase-C (PKC)-dependent mechanism. Moreover, PKC activation alone was sufficient to confer resistance to ALK inhibitors, whereas combined ALK and PKC inhibition restored sensitivity. We observed enrichment of gene signatures associated with several resistance drivers (including P2Y receptors) in crizotinib-resistant ALK-rearranged lung tumors compared to treatment-naive controls, supporting a role for these identified mechanisms in clinical ALK inhibitor resistance.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Resistencia a Medicamentos Antineoplásicos/genética , Neoplasias Pulmonares/genética , Pirazóis/farmacologia , Piridinas/farmacologia , Receptores Proteína Tirosina Quinases/antagonistas & inibidores , Quinase do Linfoma Anaplásico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Crizotinibe , Receptores ErbB/genética , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Pirazóis/uso terapêutico , Piridinas/uso terapêutico , Receptores Proteína Tirosina Quinases/genética , Receptores Proteína Tirosina Quinases/metabolismo , Receptor ErbB-2/genética , Receptores Purinérgicos P2Y/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética
16.
Proc Natl Acad Sci U S A ; 104(10): 4025-9, 2007 Mar 06.
Artigo em Inglês | MEDLINE | ID: mdl-17360471

RESUMO

The steroid hormone aldosterone is secreted both in the setting of intravascular volume depletion and hyperkalemia, raising the question of how the kidney maximizes NaCl reabsorption in the former state while maximizing K(+) secretion in the latter. Mutations in WNK4 cause pseudohypoaldosteronism type II (PHAII), a disease featuring increased renal NaCl reabsorption and impaired K(+) secretion. PHAII-mutant WNK4 achieves these effects by increasing activity of the Na-Cl cotransporter (NCC) and the Na(+) channel ENaC while concurrently inhibiting the renal outer medullary K(+) channel (ROMK). We now describe a functional state for WNK4 that promotes increased, rather than decreased, K(+) secretion. We show that WNK4 is phosphorylated by SGK1, a mediator of aldosterone signaling. Whereas wild-type WNK4 inhibits the activity of both ENaC and ROMK, a WNK4 mutation that mimics phosphorylation at the SGK1 site (WNK4(S1169D)) alleviates inhibition of both channels. The net result of these effects in the kidney would be increased K(+) secretion, because of both increased electrogenic Na(+) reabsorption and increased apical membrane K(+) permeability. Thus, modification at the PHAII and SGK1 sites in WNK4 impart opposite effects on K(+) secretion, decreasing or increasing ROMK activity and net K(+) secretion, respectively. This functional state for WNK4 would thus promote the desired physiologic response to hyperkalemia, and the fact that it is induced downstream of aldosterone signaling implicates WNK4 in the physiologic response to aldosterone with hyperkalemia. Together, the different states of WNK4 allow the kidney to provide distinct and appropriate integrated responses to intravascular volume depletion and hyperkalemia.


Assuntos
Proteínas Imediatamente Precoces/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Aldosterona/metabolismo , Sequência de Aminoácidos , Animais , Humanos , Camundongos , Dados de Sequência Molecular , Oócitos/metabolismo , Potássio/metabolismo , Canais de Potássio/metabolismo , Proteínas Serina-Treonina Quinases/química , Transdução de Sinais , Canais de Sódio/metabolismo , Xenopus laevis
17.
Proc Natl Acad Sci U S A ; 104(10): 4020-4, 2007 Mar 06.
Artigo em Inglês | MEDLINE | ID: mdl-17360470

RESUMO

Homeostasis of intravascular volume, Na(+), Cl(-), and K(+) is interdependent and determined by the coordinated activities of structurally diverse mediators in the distal nephron and the distal colon. The behavior of these flux pathways is regulated by the renin-angiotensin-aldosterone system; however, the mechanisms that allow independent modulation of individual elements have been obscure. Previous work has shown that mutations in WNK4 cause pseudohypoaldosteronism type II (PHAII), a disease featuring hypertension with hyperkalemia, due to altered activity of specific Na-Cl cotransporters, K(+) channels, and paracellular Cl(-) flux mediators of the distal nephron. By coexpression studies in Xenopus oocytes, we now demonstrate that WNK4 also inhibits the epithelial Na(+) channel (ENaC), the major mediator of aldosterone-sensitive Na(+) (re)absorption, via a mechanism that is independent of WNK4's kinase activity. This inhibition requires intact C termini in ENaC beta- and gamma-subunits, which contain PY motifs used to target ENaC for clearance from the plasma membrane. Importantly, PHAII-causing mutations eliminate WNK4's inhibition of ENaC, thereby paralleling other effects of PHAII to increase sodium balance. The relevance of these findings in vivo was studied in mice harboring PHAII-mutant WNK4. The colonic epithelium of these mice demonstrates markedly increased amiloride-sensitive Na(+) flux compared with wild-type littermates. These studies identify ENaC as a previously unrecognized downstream target of WNK4 and demonstrate a functional role for WNK4 in the regulation of colonic Na(+) absorption. These findings support a key role for WNK4 in coordinating the activities of diverse flux pathways to achieve integrated fluid and electrolyte homeostasis.


Assuntos
Canais Epiteliais de Sódio/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/fisiologia , Aldosterona/metabolismo , Animais , Colo/metabolismo , Hiperpotassemia/metabolismo , Hipertensão/metabolismo , Camundongos , Camundongos Transgênicos , Mutação , Oócitos/metabolismo , Pseudo-Hipoaldosteronismo/metabolismo , Ratos , Sistema Renina-Angiotensina , Xenopus laevis/metabolismo
18.
J Physiol ; 571(Pt 2): 275-86, 2006 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-16357011

RESUMO

The serine-threonine kinase WNK3 modulates Cl- transport into and out of cells through its regulation of SLC12A cation/Cl- cotransporters, implicating it as (one of) the long-sought Cl-/volume-sensitive kinase(s). Integrators in homeostatic systems regulate structurally diverse but functionally coupled elements. For example, the related kinase WNK4 regulates the Na-Cl co-transporter (NCC), paracellular Cl- flux, and the K+ channel ROMK1 (Kir1.1) to maintain renal NaCl and K+ homeostasis; mutations in PRKWNK4, encoding WNK4, cause a Mendelian disease featuring hypertension and hyperkalemia. It is known that WNK3 is expressed in the nephron's distal convoluted tubule (DCT) and stimulates NCC activity. Here, we show that WNK3 is also expressed in cortical and outer medullary collecting duct principal cells. Accordingly, we tested WNK3's effect on the mediators of NaCl and K+ handling in these nephron segments--the epithelial sodium channel (ENaC), paracellular Cl- flux, and ROMK1--using established model systems. WNK3 did not alter paracellular Cl- flux in tetracycline-responsive MDCK II cells, nor affect amiloride-sensitive currents when co-expressed with ENaC in Xenopus laevis oocytes. However, additional co-expression studies in oocytes revealed WNK3 inhibited the renal-specific K+ channel ROMK1 activity greater than 5.5-fold (p < .0001) by altering its plasmalemmal surface expression; WNK3 did not affect ROMK1's conductance or open/closed probability. In contrast, WNK3 had no effect on the activity of the cardiac long-QT syndrome K+ channel KCNQ1/KCNE1 when co-expressed in oocytes. Inhibition of ROMK1 is independent of WNK3's catalytic activity and is mediated by WNK3's carboxyl terminus--a mechanism distinct from its known kinase-dependent activation of NCC. A kinase-inactivating point mutation, or a missense mutation homologous to one in WNK4 that causes disease produced a gain-of-function effect, enhancing WNK3's inhibition of ROMK1 greater than 2.5-fold relative to wild type kinase (p < .0001). The magnitude and specificity of WNK3's effects at both NCC and ROMK1, its co-expression with its targets in the distal nephron, and the established in vivo effect of WNK4 at these same targets provide evidence that WNK3's action is physiologically relevant. WNK3 is likely a component of one of the mechanisms that determines the balance between renal NaCl reabsorption and K+ secretion.


Assuntos
Hiperpotassemia/genética , Hipertensão/genética , Túbulos Renais Coletores/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Proteínas Serina-Treonina Quinases/fisiologia , Animais , Linhagem Celular , Cães , Córtex Renal/metabolismo , Medula Renal/metabolismo , Camundongos , Modelos Biológicos , Mutação , Oócitos/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização/fisiologia , Proteínas Serina-Treonina Quinases/genética , Canais de Sódio/metabolismo , Simportadores de Cloreto de Sódio/metabolismo , Xenopus laevis
19.
Proc Natl Acad Sci U S A ; 102(46): 16777-82, 2005 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-16275913

RESUMO

WNK1 and WNK4 [WNK, with no lysine (K)] are serine-threonine kinases that function as molecular switches, eliciting coordinated effects on diverse ion transport pathways to maintain homeostasis during physiological perturbation. Gain-of-function mutations in either of these genes cause an inherited syndrome featuring hypertension and hyperkalemia due to increased renal NaCl reabsorption and decreased K(+) secretion. Here, we reveal unique biochemical and functional properties of WNK3, a related member of the WNK kinase family. Unlike WNK1 and WNK4, WNK3 is expressed throughout the nephron, predominantly at intercellular junctions. Because WNK4 is a potent inhibitor of members of the cation-cotransporter SLC12A family, we used coexpression studies in Xenopus oocytes to investigate the effect of WNK3 on NCC and NKCC2, related kidney-specific transporters that mediate apical NaCl reabsorption in the thick ascending limb and distal convoluted tubule, respectively. In contrast to WNK4's inhibitory activity, kinase-active WNK3 is a potent activator of both NKCC2 and NCC-mediated transport. Conversely, in its kinase-inactive state, WNK3 is a potent inhibitor of NKCC2 and NCC activity. WNK3 regulates the activity of these transporters by altering their expression at the plasma membrane. Wild-type WNK3 increases and kinase-inactive WNK3 decreases NKCC2 phosphorylation at Thr-184 and Thr-189, sites required for the vasopressin-mediated plasmalemmal translocation and activation of NKCC2 in vivo. The effects of WNK3 on these transporters and their coexpression in renal epithelia implicate WNK3 in NaCl, water, and blood pressure homeostasis, perhaps via signaling downstream of vasopressin.


Assuntos
Pressão Sanguínea/fisiologia , Homeostase , Rim/fisiologia , Proteínas Serina-Treonina Quinases/fisiologia , Simportadores de Cloreto de Sódio-Potássio/fisiologia , Simportadores/fisiologia , Animais , Células COS , Chlorocebus aethiops , Eletroforese em Gel de Poliacrilamida , Camundongos , Microscopia de Fluorescência , Fosforilação , Membro 1 da Família 12 de Carreador de Soluto , Transfecção
20.
Curr Opin Nephrol Hypertens ; 13(5): 557-62, 2004 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-15300163

RESUMO

PURPOSE OF REVIEW: The WNK kinases are a recently discovered family of serine-threonine kinases that have been shown to play an essential role in the regulation of electrolyte homeostasis. This review focuses on the recent evidence elucidating the functions of these kinases in normal and disease physiology. RECENT FINDINGS: Mutations in WNK1 and WNK4 have been shown to cause pseudohypoaldosteronism type II, a disease featuring hypertension with hyperkalemia. Recent work has demonstrated that WNK4 is a potent inhibitor of diverse epithelial transporters including the thiazide-sensitive sodium chloride co-transporter (NCCT) and the renal outer medullary potassium ion channel. In addition, WNK4 activity promotes paracellular chloride ion flux. Importantly, mutations in WNK4 that cause disease have divergent effects on these transport pathways. WNK4 mutations relieve the inhibition of NCCT, increase the inhibition of the renal outer medullary potassium ion channel, and further increase paracellular chloride ion flux. These findings can explain the observed physiological abnormalities in patients with pseudohypoaldosteronism type II, and support a model in which WNK4 is a molecular switch that can alter the balance between chloride ion reabsorption and potassium ion secretion. The WNK kinases are also found in diverse epithelia throughout the body that are involved in chloride ion flux, suggesting that these kinases may play a general role in the regulation of chloride ion flux. SUMMARY: The WNK kinases define a previously unrecognized signaling pathway that is essential for the integrated regulation of electrolyte homeostasis. Their function has implications for understanding the coordinated regulation of electrolyte homeostasis and blood pressure, and identifies WNKs as dynamic regulators of the paracellular flux pathway.


Assuntos
Proteínas Serina-Treonina Quinases/fisiologia , Epitélio/metabolismo , Humanos , Transporte de Íons/genética , Rim/metabolismo , Mutação Puntual/genética , Proteínas Serina-Treonina Quinases/classificação , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Pseudo-Hipoaldosteronismo/genética , Pseudo-Hipoaldosteronismo/metabolismo , Transdução de Sinais/fisiologia
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA