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1.
Nat Commun ; 12(1): 5395, 2021 09 13.
Artigo em Inglês | MEDLINE | ID: mdl-34518531

RESUMO

Knowledge of the genomic landscape of chronic lymphocytic leukemia (CLL) grows increasingly detailed, providing challenges in contextualizing the accumulated information. To define the underlying networks, we here perform a multi-platform molecular characterization. We identify major subgroups characterized by genomic instability (GI) or activation of epithelial-mesenchymal-transition (EMT)-like programs, which subdivide into non-inflammatory and inflammatory subtypes. GI CLL exhibit disruption of genome integrity, DNA-damage response and are associated with mutagenesis mediated through activation-induced cytidine deaminase or defective mismatch repair. TP53 wild-type and mutated/deleted cases constitute a transcriptionally uniform entity in GI CLL and show similarly poor progression-free survival at relapse. EMT-like CLL exhibit high genomic stability, reduced benefit from the addition of rituximab and EMT-like differentiation is inhibited by induction of DNA damage. This work extends the perspective on CLL biology and risk categories in TP53 wild-type CLL. Furthermore, molecular targets identified within each subgroup provide opportunities for new treatment approaches.


Assuntos
Transição Epitelial-Mesenquimal/genética , Perfilação da Expressão Gênica/métodos , Regulação Leucêmica da Expressão Gênica , Redes Reguladoras de Genes , Instabilidade Genômica , Leucemia Linfocítica Crônica de Células B/genética , Proteínas Mutadas de Ataxia Telangiectasia/genética , Aberrações Cromossômicas , Dano ao DNA , Reparo do DNA , Humanos , Mutação , Polimorfismo de Nucleotídeo Único , Complexo Shelterina , Proteínas de Ligação a Telômeros/genética , Proteína Supressora de Tumor p53/genética
2.
Invest New Drugs ; 38(3): 844-854, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-31385109

RESUMO

Background Endothelin B receptor (ETBR) is involved in melanoma pathogenesis and is overexpressed in metastatic melanoma. The antibody-drug conjugate DEDN6526A targets ETBR and is comprised of the humanized anti-ETBR monoclonal antibody conjugated to the anti-mitotic agent monomethyl auristatin E (MMAE). Methods This Phase I study evaluated the safety, pharmacokinetics, pharmacodynamics, and anti-tumor activity of DEDN6526A (0.3-2.8 mg/kg) given every 3 weeks (q3w) in patients with metastatic or unresectable cutaneous, mucosal, or uveal melanoma. Results Fifty-three patients received a median of 6 doses of DEDN6526A (range 1-49). The most common drug-related adverse events (>25% across dose levels) were fatigue, peripheral neuropathy, nausea, diarrhea, alopecia, and chills. Three patients in dose-escalation experienced a dose-limiting toxicity (infusion-related reaction, increased ALT/AST, and drug-induced liver injury). Based on cumulative safety data across all dose levels, the recommended Phase II dose (RP2D) for DEDN6526A was 2.4 mg/kg intravenous (IV) q3w. The pharmacokinetics of antibody-conjugated MMAE and total antibody were dose-proportional at doses ranging from 1.8-2.8 mg/kg. A trend toward faster clearance was observed at doses of 0.3-1.2 mg/kg. There were 6 partial responses (11%) in patients with metastatic cutaneous or mucosal melanoma, and 17 patients (32%) had prolonged stable disease ≥6 months. Responses were independent of BRAF mutation status but did correlate with ETBR expression. Conclusion DEDN6526A administered at the RP2D of 2.4 mg/kg q3w had an acceptable safety profile and showed evidence of anti-tumor activity in patients with cutaneous, mucosal, and uveal melanoma. ClinicalTrials.gov identifier: NCT01522664.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Antagonistas do Receptor de Endotelina B/uso terapêutico , Imunoconjugados/uso terapêutico , Melanoma/tratamento farmacológico , Receptor de Endotelina B/metabolismo , Neoplasias Uveais/tratamento farmacológico , Adulto , Idoso , Idoso de 80 Anos ou mais , Esquema de Medicação , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Adulto Jovem
3.
Clin Cancer Res ; 22(12): 2874-84, 2016 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-26787751

RESUMO

PURPOSE: This first-in-human phase I trial assessed the safety, tolerability, and preliminary antitumor activity of apitolisib (GDC-0980), a dual inhibitor of class I PI3K, and mTOR kinases. EXPERIMENTAL DESIGN: Once-daily oral apitolisib was administered to patients with solid tumors for days 1 to 21 or 1 to 28 of 28-day cycles. Pharmacokinetic and pharmacodynamic parameters were assessed. RESULTS: Overall, 120 patients were treated at doses between 2 and 70 mg. The commonest ≥G3 toxicities related to apitolisib at the recommended phase 2 dose (RP2D) at 40 mg once daily included hyperglycemia (18%), rash (14%), liver dysfunction (12%), diarrhea (10%), pneumonitis (8%), mucosal inflammation (6%), and fatigue (4%). Dose-limiting toxicities (1 patient each) were G4 fasting hyperglycemia at 40 mg (21/28 schedule) and G3 maculopapular rash and G3 fasting hyperglycemia at 70 mg (21/28 schedule). The pharmacokinetic profile was dose-proportional. Phosphorylated serine-473 AKT levels were suppressed by ≥90% in platelet-rich plasma within 4 hours at the MTD (50 mg). Pharmacodynamic decreases in fluorodeoxyglucose positron emission tomography uptake of >25% occurred in 66% (21/32) of patients dosed at 40 mg once daily. Evidence of single-agent activity included 10 RECIST partial responses (PR; confirmed for peritoneal mesothelioma, PIK3CA mutant head-and-neck cancer, and three pleural mesotheliomas). CONCLUSIONS: Apitolisib exhibited dose-proportional pharmacokinetics with target modulation at doses ≥16 mg. The RP2D was 40 mg once-daily 28/28 schedule; severe on-target toxicities were apparent at ≥40 mg, particularly pneumonitis. Apitolisib was reasonably tolerated at 30 mg, the selected dose for pleural mesothelioma patients given limited respiratory reserve. Modest but durable antitumor activity was demonstrated. Clin Cancer Res; 22(12); 2874-84. ©2016 AACR.


Assuntos
Antineoplásicos/uso terapêutico , Compostos Bicíclicos Heterocíclicos com Pontes/uso terapêutico , Classe I de Fosfatidilinositol 3-Quinases/antagonistas & inibidores , Neoplasias/tratamento farmacológico , Inibidores de Proteínas Quinases/uso terapêutico , Pirimidinas/uso terapêutico , Serina-Treonina Quinases TOR/antagonistas & inibidores , Antineoplásicos/efeitos adversos , Antineoplásicos/farmacocinética , Compostos Bicíclicos Heterocíclicos com Pontes/efeitos adversos , Compostos Bicíclicos Heterocíclicos com Pontes/farmacocinética , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Inibidores de Proteínas Quinases/efeitos adversos , Inibidores de Proteínas Quinases/farmacocinética , Pirimidinas/efeitos adversos , Pirimidinas/farmacocinética
4.
Blood ; 124(3): 420-5, 2014 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-24916506

RESUMO

Addition of rituximab (R) to fludarabine and cyclophosphamide (FC) has significantly improved patient outcomes in chronic lymphocytic leukemia (CLL). Whether baseline gene expression can identify patients who will benefit from immunochemotherapy over chemotherapy alone has not been determined. We assessed genome-wide expression of 300 pretreatment specimens from a subset of 552 patients in REACH, a study of FC or R-FC in relapsed CLL. An independent test set was derived from 282 pretreatment specimens from CLL8, a study of FC or R-FC in treatment-naïve patients. Genes specific for benefit from R-FC were determined by assessing treatment-gene interactions in Cox proportional hazards models. REACH patients with higher pretreatment protein tyrosine kinase 2 (PTK2) messenger RNA levels derived greater benefit from R-FC, with significant improvements in progression-free survival, independent of known prognostic factors in a multivariate model. Examination of PTK2 gene expression in CLL8 patients yielded similar results. Furthermore, PTK2 inhibition blunted R-dependent cell death in vitro. This retrospective analysis from 2 independent trials revealed that increased PTK2 expression is associated with improved outcomes for CLL patients treated with R-FC vs FC. PTK2 expression may be a useful biomarker for patient selection in future trials. These trials were registered at www.clinicaltrials.gov as #NCT00090051 (REACH) and #NCT00281918 (CLL8).


Assuntos
Quinase 1 de Adesão Focal/genética , Leucemia Linfocítica Crônica de Células B/genética , Leucemia Linfocítica Crônica de Células B/terapia , Anticorpos Monoclonais Murinos/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Terapia Combinada , Ciclofosfamida/administração & dosagem , Intervalo Livre de Doença , Expressão Gênica , Humanos , Imunoterapia , Leucemia Linfocítica Crônica de Células B/enzimologia , Modelos de Riscos Proporcionais , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Neoplásico/genética , RNA Neoplásico/metabolismo , Recidiva , Estudos Retrospectivos , Rituximab , Vidarabina/administração & dosagem , Vidarabina/análogos & derivados
5.
Contemp Clin Trials ; 36(2): 624-35, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-23648396

RESUMO

In the past decade, the cost of drug development has increased significantly. The estimates vary widely but frequently quoted numbers are staggering-it takes 10-15 years and billions of dollars to bring a drug to patients. To a large extent this is due to many long, expensive and ultimately unsuccessful drug trials. While one approach to combat the low yield on investment could be to continue searching for new blockbusters, an alternative method would lead us to focus on testing new targeted treatments that have a strong underlying scientific rationale and are more likely to provide enhanced clinical benefit in population subsets defined by molecular diagnostics. Development of these new treatments, however, cannot follow the usual established path; new strategies and approaches are required for the co-development of novel therapeutics and the diagnostic. In this paper we will review, from the point of view of industry, the approaches to, and challenges of drug development strategies incorporating predictive biomarkers into clinical programs. We will outline the basic concepts behind co-development with predictive biomarkers and summarize the current regulatory paradigm. We will present guiding principles of personalized health care (PHC) development and review the statistical, strategic, regulatory and operational challenges that statisticians regularly encounter on development programs with a PHC component. Some practical recommendations for team statisticians involved in PHC drug development are included. The majority of the examples and recommendations are drawn from oncology but broader concepts apply across all therapeutic areas.


Assuntos
Aprovação de Drogas , Indústria Farmacêutica , Medicina de Precisão , Biomarcadores/metabolismo , Ensaios Clínicos Fase III como Assunto/métodos , Ensaios Clínicos Fase III como Assunto/normas , Interpretação Estatística de Dados , Aprovação de Drogas/métodos , Indústria Farmacêutica/métodos , Tratamento Farmacológico , Humanos , Medicina de Precisão/métodos , Tamanho da Amostra
6.
Clin Cancer Res ; 19(13): 3681-92, 2013 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-23685835

RESUMO

PURPOSE: The aim of this study was to identify conserved pharmacodynamic and potential predictive biomarkers of response to anti-VEGF therapy using gene expression profiling in preclinical tumor models and in patients. EXPERIMENTAL DESIGN: Surrogate markers of VEGF inhibition [VEGF-dependent genes or VEGF-dependent vasculature (VDV)] were identified by profiling gene expression changes induced in response to VEGF blockade in preclinical tumor models and in human biopsies from patients treated with anti-VEGF monoclonal antibodies. The potential value of VDV genes as candidate predictive biomarkers was tested by correlating high or low VDV gene expression levels in pretreatment clinical samples with the subsequent clinical efficacy of bevacizumab (anti-VEGF)-containing therapy. RESULTS: We show that VDV genes, including direct and more distal VEGF downstream endothelial targets, enable detection of VEGF signaling inhibition in mouse tumor models and human tumor biopsies. Retrospective analyses of clinical trial data indicate that patients with higher VDV expression in pretreatment tumor samples exhibited improved clinical outcome when treated with bevacizumab-containing therapies. CONCLUSIONS: In this work, we identified surrogate markers (VDV genes) for in vivo VEGF signaling in tumors and showed clinical data supporting a correlation between pretreatment VEGF bioactivity and the subsequent efficacy of anti-VEGF therapy. We propose that VDV genes are candidate biomarkers with the potential to aid the selection of novel indications as well as patients likely to respond to anti-VEGF therapy. The data presented here define a diagnostic biomarker hypothesis based on translational research that warrants further evaluation in additional retrospective and prospective trials.


Assuntos
Inibidores da Angiogênese/uso terapêutico , Antineoplásicos/uso terapêutico , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Fator A de Crescimento do Endotélio Vascular/metabolismo , Inibidores da Angiogênese/farmacologia , Animais , Anticorpos Monoclonais Humanizados/farmacologia , Anticorpos Monoclonais Humanizados/uso terapêutico , Antineoplásicos/farmacologia , Bevacizumab , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Camundongos , Neoplasias/genética , Neoplasias/mortalidade , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Tumores Neuroendócrinos/tratamento farmacológico , Tumores Neuroendócrinos/genética , Tumores Neuroendócrinos/metabolismo , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo
7.
Blood ; 121(18): 3650-7, 2013 May 02.
Artigo em Inglês | MEDLINE | ID: mdl-23525797

RESUMO

In chronic lymphocytic leukemia (CLL) patients, disruptions of the TP53 tumor suppressor pathway by 17p13 deletion (del17p), somatic TP53 mutations, or downregulation of microRNA-34a have been associated with a poor prognosis. So far, the impact of the various TP53 defects has not been evaluated in a large cohort of previously treated and relapsed CLL patients. Here, we present the results of TP53 gene sequencing and fluorescence in situ hybridization for del17p in a phase 3 clinical trial (REACH [Rituximab in the Study of Relapsed Chronic Lymphocytic Leukemia]). Of the 457 patients, 52 had TP53 mutations and 37 had del17p. In 24 (46%) of the TP53 mutated patients, no del17p was found and in 9 of the del17p patients, no TP53 mutation was identified. Based on a predicted proportion of TP53 disruption, a complete disruption of TP53 function, either by a combination of point mutations and/or del17p, was associated with a high risk for disease progression. Progression-free survival of patients with a heterozygous TP53 mutation was not significantly different from patients with a completely intact TP53 locus. In addition, only a complete loss of TP53 function correlated with low microRNA-34a expression levels. This trial was registered at www.clinicaltrials.gov as #NCT00090051.


Assuntos
Leucemia Linfocítica Crônica de Células B/genética , MicroRNAs/genética , Proteína Supressora de Tumor p53/genética , Adulto , Idoso , Biomarcadores Tumorais/genética , Progressão da Doença , Intervalo Livre de Doença , Regulação para Baixo/genética , Feminino , Regulação Leucêmica da Expressão Gênica , Inativação Gênica/fisiologia , Humanos , Leucemia Linfocítica Crônica de Células B/diagnóstico , Leucemia Linfocítica Crônica de Células B/patologia , Leucemia Linfocítica Crônica de Células B/terapia , Masculino , Pessoa de Meia-Idade , Prognóstico , Recidiva , Falha de Tratamento
8.
Hum Mol Genet ; 21(9): 2054-67, 2012 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-22286172

RESUMO

Pluripotent stem cells are derived from culture of early embryos or the germline and can be induced by reprogramming of somatic cells. Barriers to reprogramming that stabilize the differentiated state and have tumor suppression functions are expected to exist. However, we have a limited understanding of what such barriers might be. To find novel barriers to reprogramming to pluripotency, we compared the transcriptional profiles of the mouse germline with pluripotent and somatic cells, in vivo and in vitro. There is a remarkable global expression of the transcriptional program for pluripotency in primordial germ cells (PGCs). We identify parallels between PGC reprogramming to pluripotency and human germ cell tumorigenesis, including the loss of LATS2, a tumor suppressor kinase of the Hippo pathway. We show that knockdown of LATS2 increases the efficiency of induction of pluripotency in human cells. LATS2 RNAi, unlike p53 RNAi, specifically enhances the generation of fully reprogrammed iPS cells without accelerating cell proliferation. We further show that LATS2 represses reprogramming in human cells by post-transcriptionally antagonizing TAZ but not YAP, two downstream effectors of the Hippo pathway. These results reveal transcriptional parallels between germ cell transformation and the generation of iPS cells and indicate that the Hippo pathway constitutes a barrier to cellular reprogramming.


Assuntos
Reprogramação Celular/genética , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo , Aciltransferases , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proteínas de Ciclo Celular , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Perfilação da Expressão Gênica , Técnicas de Silenciamento de Genes , Genes p53 , Células Germinativas/citologia , Células Germinativas/metabolismo , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Camundongos , Camundongos Transgênicos , Modelos Biológicos , Neoplasias Embrionárias de Células Germinativas/etiologia , Neoplasias Embrionárias de Células Germinativas/genética , Neoplasias Embrionárias de Células Germinativas/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Interferência de RNA , Transdução de Sinais , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas Supressoras de Tumor/antagonistas & inibidores , Proteínas de Sinalização YAP
9.
Sci Signal ; 4(186): pt5, 2011 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-21868360

RESUMO

Compared with the luminal subtype, the basal-like subtype of breast cancer has an aggressive clinical behavior, but the reasons for this difference between the two subtypes are poorly understood. We identified microRNAs (miRNAs) miR-221 and miR-222 (miR-221/222) as basal-like subtype-specific miRNAs that decrease expression of epithelial-specific genes and increase expression of mesenchymal-specific genes. In addition, expression of these miRNAs increased cell migration and invasion, which collectively are characteristics of the epithelial-to-mesenchymal transition (EMT). The basal-like transcription factor FOSL1 (also known as Fra-1) directly stimulated the transcription of miR-221/222, and the abundance of these miRNAs decreased with inhibition of MEK (mitogen-activated or extracellular signal-regulated protein kinase kinase), placing miR-221/222 downstream of the RAS pathway. The miR-221/222-mediated reduction in E-cadherin abundance depended on their targeting of the 3' untranslated region (3'UTR) of TRPS1 (trichorhinophalangeal syndrome type 1), which is a member of the GATA family of transcriptional repressors. TRPS1 inhibited EMT by directly repressing expression of ZEB2 (Zinc finger E-box-binding homeobox 2). Therefore, miR-221/222 may contribute to the aggressive clinical behavior of basal-like breast cancers.


Assuntos
Neoplasias da Mama/metabolismo , Proteínas de Ligação a DNA/biossíntese , Transição Epitelial-Mesenquimal , MicroRNAs/metabolismo , Proteínas de Neoplasias/metabolismo , RNA Neoplásico/metabolismo , Fatores de Transcrição/biossíntese , Regiões 3' não Traduzidas/genética , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Movimento Celular/genética , Proteínas de Ligação a DNA/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , MAP Quinase Quinase Quinases/genética , MAP Quinase Quinase Quinases/metabolismo , MicroRNAs/genética , Invasividade Neoplásica , Proteínas de Neoplasias/genética , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Proto-Oncogênicas c-fos/metabolismo , RNA Neoplásico/genética , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Fatores de Transcrição/genética , Homeobox 2 de Ligação a E-box com Dedos de Zinco
10.
Sci Signal ; 4(177): ra41, 2011 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-21673316

RESUMO

The basal-like subtype of breast cancer has an aggressive clinical behavior compared to that of the luminal subtype. We identified the microRNAs (miRNAs) miR-221 and miR-222 (miR-221/222) as basal-like subtype-specific miRNAs and showed that expression of miR-221/222 decreased expression of epithelial-specific genes and increased expression of mesenchymal-specific genes, and increased cell migration and invasion in a manner characteristic of the epithelial-to-mesenchymal transition (EMT). The transcription factor FOSL1 (also known as Fra-1), which is found in basal-like breast cancers but not in the luminal subtype, stimulated the transcription of miR-221/222, and the abundance of these miRNAs decreased with inhibition of the epidermal growth factor receptor (EGFR) or MEK (mitogen-activated or extracellular signal-regulated protein kinase kinase), placing miR-221/222 downstream of the RAS pathway. Furthermore, miR-221/222-mediated reduction in E-cadherin abundance depended on their targeting the 3' untranslated region of the GATA family transcriptional repressor TRPS1 (tricho-rhino-phalangeal syndrome type 1), which inhibited EMT by decreasing ZEB2 (zinc finger E-box-binding homeobox2) expression. We conclude that by promoting EMT, miR-221/222 may contribute to the more aggressive clinical behavior of basal-like breast cancers.


Assuntos
Neoplasias da Mama/metabolismo , Proteínas de Ligação a DNA/biossíntese , Transição Epitelial-Mesenquimal , Regulação Neoplásica da Expressão Gênica , MicroRNAs/biossíntese , RNA Neoplásico/biossíntese , Fatores de Transcrição/biossíntese , Regiões 3' não Traduzidas/genética , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Receptores ErbB/genética , Receptores ErbB/metabolismo , Feminino , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , MAP Quinase Quinase Quinases/genética , MAP Quinase Quinase Quinases/metabolismo , MicroRNAs/genética , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Proto-Oncogênicas c-fos/metabolismo , RNA Neoplásico/genética , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Fatores de Transcrição/genética , Homeobox 2 de Ligação a E-box com Dedos de Zinco , Proteínas ras/genética , Proteínas ras/metabolismo
11.
PLoS One ; 5(9): e12517, 2010 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-20838621

RESUMO

BACKGROUND: Evaluation of cancer biomarkers from blood could significantly enable biomarker assessment by providing a relatively non-invasive source of representative tumor material. Circulating Tumor Cells (CTCs) isolated from blood of metastatic cancer patients hold significant promise in this regard. METHODOLOGY/PRINCIPAL FINDINGS: Using spiked tumor-cells we evaluated CTC capture on different CTC technology platforms, including CellSearch and two biochip platforms, and used the isolated CTCs to develop and optimize assays for molecular characterization of CTCs. We report similar performance for the various platforms tested in capturing CTCs, and find that capture efficiency is dependent on the level of EpCAM expression. We demonstrate that captured CTCs are amenable to biomarker analyses such as HER2 status, qRT-PCR for breast cancer subtype markers, KRAS mutation detection, and EGFR staining by immunofluorescence (IF). We quantify cell surface expression of EGFR in metastatic lung cancer patient samples. In addition, we determined HER2 status by IF and FISH in CTCs from metastatic breast cancer patients. In the majority of patients (89%) we found concordance with HER2 status from patient tumor tissue, though in a subset of patients (11%), HER2 status in CTCs differed from that observed in the primary tumor. Surprisingly, we found CTC counts to be higher in ER+ patients in comparison to HER2+ and triple negative patients, which could be explained by low EpCAM expression and a more mesenchymal phenotype of tumors belonging to the basal-like molecular subtype of breast cancer. CONCLUSIONS/SIGNIFICANCE: Our data suggests that molecular characterization from captured CTCs is possible and can potentially provide real-time information on biomarker status. In this regard, CTCs hold significant promise as a source of tumor material to facilitate clinical biomarker evaluation. However, limitations exist from a purely EpCAM based capture system and addition of antibodies to mesenchymal markers could further improve CTC capture efficiency to enable routine biomarker analysis from CTCs.


Assuntos
Regulação Neoplásica da Expressão Gênica , Células Neoplásicas Circulantes/metabolismo , Receptor ErbB-2/genética , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Feminino , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Masculino , Receptor ErbB-2/metabolismo
12.
Blood ; 116(23): 4885-93, 2010 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-20807887

RESUMO

There is increasing evidence that miRNA and transcription factors interact in an instructive fashion in normal and malignant hematopoiesis. We explored the impact of TEL-AML1 (ETV6-RUNX1), the most common fusion protein in childhood leukemia, on miRNA expression and the leukemic phenotype. Using RNA interference, miRNA expression arrays, and quantitative polymerase chain reaction, we identified miRNA-494 and miRNA-320a to be up-regulated upon TEL-AML1 silencing independently of TEL expression. Chromatin immunoprecipitation analysis identified miRNA-494 as a direct miRNA target of the fusion protein TEL-AML1. Using bioinformatic analysis as well as functional luciferase experiments, we demonstrate that survivin is a target of the 2 miRNAs. miRNA-494 and miRNA-320a were introduced to the cells by transfection and survivin expression determined by Western blot analysis. These miRNAs blocked survivin expression and resulted in apoptosis in a similar manner as TEL-AML1 silencing by itself; this silencing was also shown to be Dicer-dependent. miRNAs-494 and -320a are expressed at lower levels in TEL-AML1+ leukemias compared with immunophenotype-matched nonTEL-AML1 acute lymphoblastic leukemia subtypes, and within TEL-AML1+ leukemias their expression is correlated to survivin levels. In summary our data suggest that TEL-AML1 might exert its antiapoptotic action at least in part by suppressing miRNA-494 and miRNA-320a, lowering their expression causing enhanced survivin expression.


Assuntos
Apoptose/genética , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Regulação Neoplásica da Expressão Gênica , MicroRNAs/genética , Proteínas Associadas aos Microtúbulos/genética , Proteínas de Fusão Oncogênica/genética , Adolescente , Western Blotting , Linhagem Celular Tumoral , Criança , Pré-Escolar , Imunoprecipitação da Cromatina , Expressão Gênica , Humanos , Proteínas Inibidoras de Apoptose , Proteínas Associadas aos Microtúbulos/biossíntese , Análise de Sequência com Séries de Oligonucleotídeos , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , RNA Mensageiro/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Survivina , Transfecção
13.
Blood ; 116(20): 4212-22, 2010 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-20705761

RESUMO

Polymorphisms of activating Fc-γ receptors (FCGRs) on natural killer cells and macrophages result in variable affinity for immunoglobulin G1 monoclonal antibodies and subsequently modulate antibody-dependent cellular cytotoxicity (ADCC) activity. Whether single-nucleotide polymorphisms of FCGRs correlate with survival of chronic lymphocytic leukemia (CLL) patients treated with a monoclonal antibody containing regimen is unclear. We assessed the FCGR3A and FCGR2A genotype of patients enrolled in the REACH trial, where patients received fludarabine and cyclophosphamide (FC) or rituximab plus FC (R-FC). FCGR3A and FCGR2A polymorphisms did not demonstrate prognostic significance in the FC arm (P = .42 and P = .64, respectively) or R-FC arm (P = .41 and P = .88, respectively) with respect to progression free survival. Patients with intermediate affinity genotypes (FV and HR) benefited significantly from addition of rituximab (hazard ratio = 0.55 [0.37-0.8 CI]; P = .0017 and hazard ratio = 0.63 [0.44-0.9 CI]; P = .011, respectively). Similar benefit was suggested for patients with high- affinity VV and HH (hazard ratio = 0.86 [0.4-1.84 CI]; P = .7 and hazard ratio = 0.7 [0.41-1.18 CI]; P = .18, respectively) and low-affinity FF and RR (hazard ratio = 0.85 [0.56-1.29 CI]; P = .44 and hazard ratio = 0.82 [0.47-1.42 CI]; P = .48, respectively). Overall, our results suggest that FCGR2A and FCGR3A polymorphisms do not significantly influence the outcomes of relapsed or refractory CLL patients treated with FC or the monoclonal antibody regimen R-FC.


Assuntos
Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Polimorfismo de Nucleotídeo Único/genética , Receptores de IgG/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Monoclonais Murinos/farmacocinética , Anticorpos Monoclonais Murinos/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Ciclofosfamida/uso terapêutico , Intervalo Livre de Doença , Feminino , Genótipo , Humanos , Leucemia Linfocítica Crônica de Células B/diagnóstico , Leucemia Linfocítica Crônica de Células B/genética , Masculino , Pessoa de Meia-Idade , Prognóstico , Rituximab , Resultado do Tratamento , Vidarabina/análogos & derivados , Vidarabina/uso terapêutico
14.
Radiology ; 254(2): 564-76, 2010 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-20093527

RESUMO

PURPOSE: To determine whether magnetic resonance (MR) imaging is influenced by genetic and cellular features of glioblastoma multiforme (GBM) aggressiveness. MATERIALS AND METHODS: In this HIPAA-compliant institutional review board-approved study, multiple enhancing and peritumoral nonenhancing stereotactic neurosurgical biopsy samples from treatment-naïve GBMs were collected prospectively, with guidance from cerebral blood volume (CBV) MR imaging measurements. By using monoclonal antibodies, tissue specimens were examined for microvascular expression, hypoxia, tumor and overall cellular density, and histopathologic features of GBM aggressiveness. Genetic expression patterns were investigated with RNA microarrays. Imaging and histopathologic variables were compared with the Welch t test and Pearson correlations. Microarray analysis was performed by using false discovery rate (FDR) statistics. RESULTS: Tumor biopsy of 13 adult patients yielded 16 enhancing and 14 peritumoral nonenhancing specimens. Enhancing regions had elevated relative CBV and reduced relative apparent diffusion coefficient (ADC) measurements compared with peritumoral nonenhancing biopsy regions (P < .01). A positive correlation was found between relative CBV and all histopathologic features of aggressiveness (P < .04). An inverse correlation was found between relative ADC and all histopathologic features of aggressiveness (P < .05). RNA expression patterns between tumor regions were found to be significantly different (FDR < 0.05), with hierarchical clustering by biopsy region only. CONCLUSION: These findings suggest MR imaging is significantly influenced by GBM genetic and cellular biologic features of aggressiveness and imply physiologic MR imaging may be useful in pinpointing regions of highest malignancy within heterogeneous tissues, thus facilitating histologic grading of primary glial brain tumors.


Assuntos
Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Glioblastoma/genética , Glioblastoma/patologia , Imageamento por Ressonância Magnética/métodos , Adulto , Idoso , Biópsia , Meios de Contraste , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Imageamento Tridimensional , Modelos Lineares , Masculino , Análise em Microsséries , Pessoa de Meia-Idade , Estudos Prospectivos , RNA/metabolismo
15.
PLoS Genet ; 5(8): e1000602, 2009 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-19680444

RESUMO

Epigenetic control of gene transcription is critical for normal human development and cellular differentiation. While alterations of epigenetic marks such as DNA methylation have been linked to cancers and many other human diseases, interindividual epigenetic variations in normal tissues due to aging, environmental factors, or innate susceptibility are poorly characterized. The plasticity, tissue-specific nature, and variability of gene expression are related to epigenomic states that vary across individuals. Thus, population-based investigations are needed to further our understanding of the fundamental dynamics of normal individual epigenomes. We analyzed 217 non-pathologic human tissues from 10 anatomic sites at 1,413 autosomal CpG loci associated with 773 genes to investigate tissue-specific differences in DNA methylation and to discern how aging and exposures contribute to normal variation in methylation. Methylation profile classes derived from unsupervised modeling were significantly associated with age (P<0.0001) and were significant predictors of tissue origin (P<0.0001). In solid tissues (n = 119) we found striking, highly significant CpG island-dependent correlations between age and methylation; loci in CpG islands gained methylation with age, loci not in CpG islands lost methylation with age (P<0.001), and this pattern was consistent across tissues and in an analysis of blood-derived DNA. Our data clearly demonstrate age- and exposure-related differences in tissue-specific methylation and significant age-associated methylation patterns which are CpG island context-dependent. This work provides novel insight into the role of aging and the environment in susceptibility to diseases such as cancer and critically informs the field of epigenomics by providing evidence of epigenetic dysregulation by age-related methylation alterations. Collectively we reveal key issues to consider both in the construction of reference and disease-related epigenomes and in the interpretation of potentially pathologically important alterations.


Assuntos
Envelhecimento/genética , Ilhas de CpG , Metilação de DNA , Exposição Ambiental , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Epigênese Genética , Feminino , Humanos , Recém-Nascido , Masculino , Pessoa de Meia-Idade , Especificidade de Órgãos
16.
Cancer Res ; 69(15): 6315-21, 2009 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-19638575

RESUMO

Pathologic differentiation of tissue of origin in tumors found in the lung can be challenging, with differentiation of mesothelioma and lung adenocarcinoma emblematic of this problem. Indeed, proper classification is essential for determination of treatment regimen for these diseases, making accurate and early diagnosis critical. Here, we investigate the potential of epigenetic profiles of lung adenocarcinoma, mesothelioma, and nonmalignant pulmonary tissues (n = 285) as differentiation markers in an analysis of DNA methylation at 1413 autosomal CpG loci associated with 773 cancer-related genes. Using an unsupervised recursively partitioned mixture modeling technique for all samples, the derived methylation profile classes were significantly associated with sample type (P < 0.0001). In a similar analysis restricted to tumors, methylation profile classes significantly predicted tumor type (P < 0.0001). Random forests classification of CpG methylation of tumors--which splits the data into training and test sets--accurately differentiated mesothelioma from lung adenocarcinoma over 99% of the time (P < 0.0001). In a locus-by-locus comparison of CpG methylation between tumor types, 1266 CpG loci had significantly different methylation between tumors following correction for multiple comparisons (Q < 0.05); 61% had higher methylation in adenocarcinoma. Using the CpG loci with significant differential methylation in a pathway analysis revealed significant enrichment of methylated gene-loci in Cell Cycle Regulation, DNA Damage Response, PTEN Signaling, and Apoptosis Signaling pathways in lung adenocarcinoma when compared with mesothelioma. Methylation profile-based differentiation of lung adenocarcinoma and mesothelioma is highly accurate, informs on the distinct etiologies of these diseases, and holds promise for clinical application.


Assuntos
Adenocarcinoma/genética , Metilação de DNA , Pneumopatias/genética , Neoplasias Pulmonares/genética , Mesotelioma/genética , Neoplasias Pleurais/genética , Adenocarcinoma/diagnóstico , Adulto , Idoso , Idoso de 80 Anos ou mais , Ilhas de CpG , Citosina/metabolismo , Diagnóstico Diferencial , Feminino , Humanos , Pneumopatias/diagnóstico , Neoplasias Pulmonares/diagnóstico , Masculino , Mesotelioma/diagnóstico , Pessoa de Meia-Idade , Neoplasias Pleurais/diagnóstico
17.
Nat Genet ; 41(8): 905-8, 2009 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-19578366

RESUMO

The causes of glioblastoma and other gliomas remain obscure. To discover new candidate genes influencing glioma susceptibility, we conducted a principal component-adjusted genome-wide association study (GWAS) of 275,895 autosomal variants among 692 adult high-grade glioma cases (622 from the San Francisco Adult Glioma Study (AGS) and 70 from the Cancer Genome Atlas (TCGA)) and 3,992 controls (602 from AGS and 3,390 from Illumina iControlDB (iControls)). For replication, we analyzed the 13 SNPs with P < 10(-6) using independent data from 176 high-grade glioma cases and 174 controls from the Mayo Clinic. On 9p21, rs1412829 near CDKN2B had discovery P = 3.4 x 10(-8), replication P = 0.0038 and combined P = 1.85 x 10(-10). On 20q13.3, rs6010620 intronic to RTEL1 had discovery P = 1.5 x 10(-7), replication P = 0.00035 and combined P = 3.40 x 10(-9). For both SNPs, the direction of association was the same in discovery and replication phases.


Assuntos
Inibidor de Quinase Dependente de Ciclina p15/genética , DNA Helicases/genética , Predisposição Genética para Doença , Glioma/genética , Polimorfismo de Nucleotídeo Único/genética , Adulto , Estudos de Casos e Controles , Mapeamento Cromossômico , Cromossomos Humanos Par 9/genética , Genoma Humano/genética , Estudo de Associação Genômica Ampla , Haplótipos , Humanos , Modelos Genéticos , Análise de Componente Principal , Análise de Regressão
18.
Carcinogenesis ; 30(3): 416-22, 2009 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-19126652

RESUMO

Head and neck squamous cell carcinomas (HNSCCs) represent clinically and etiologically heterogeneous tumors affecting >40 000 patients per year in the USA. Previous research has identified individual epigenetic alterations and, in some cases, the relationship of these alterations with carcinogen exposure or patient outcomes, suggesting that specific exposures give rise to specific types of molecular alterations in HNSCCs. Here, we describe how different etiologic factors are reflected in the molecular character and clinical outcome of these tumors. In a case series of primary, incident HNSCC (n = 68), we examined the DNA methylation profile of 1413 autosomal CpG loci in 773 genes, in relation to exposures and etiologic factors. The overall pattern of epigenetic alteration could significantly distinguish tumor from normal head and neck epithelial tissues (P < 0.0001) more effectively than specific gene methylation events. Among tumors, there were significant associations between specific DNA methylation profile classes and tobacco smoking and alcohol exposures. Although there was a significant association between methylation profile and tumor stage (P < 0.01), we did not observe an association between these profiles and overall patient survival after adjustment for stage; although methylation of a number of specific loci falling in different cellular pathways was associated with overall patient survival. We found that the etiologic heterogeneity of HNSCC is reflected in specific patterns of molecular epigenetic alterations within the tumors and that the DNA methylation profiles may hold clinical promise worthy of further study.


Assuntos
Carcinoma de Células Escamosas/genética , Metilação de DNA/fisiologia , Epigênese Genética , Neoplasias de Cabeça e Pescoço/genética , Idoso , Consumo de Bebidas Alcoólicas/efeitos adversos , Biomarcadores Tumorais/metabolismo , Carcinoma de Células Escamosas/etiologia , Carcinoma de Células Escamosas/metabolismo , Ilhas de CpG , Metilação de DNA/genética , Feminino , Perfilação da Expressão Gênica/métodos , Neoplasias de Cabeça e Pescoço/etiologia , Neoplasias de Cabeça e Pescoço/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Fumar/efeitos adversos
19.
Cancer Res ; 69(1): 227-34, 2009 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-19118007

RESUMO

Mechanisms of action of nonmutagenic carcinogens such as asbestos remain poorly characterized. As pleural mesothelioma is known to have limited numbers of genetic mutations, we aimed to characterize the relationships among gene-locus-specific methylation alterations, disease status, asbestos burden, and survival in this rapidly fatal asbestos-associated tumor. Methylation of 1505 CpG loci associated with 803 cancer-related genes were studied in 158 pleural mesotheliomas and 18 normal pleura. After false-discovery rate correction, 969 CpG loci were independently associated with disease status (Q < 0.05). Classifying samples based on CpG methylation profile with a mixture model approach, methylation classes discriminated tumor from normal pleura (permutation P < 0.0001). In a random forests classification, the overall misclassification error rate was 3.4%, with <1% (n = 1) of tumors misclassified as normal (P < 0.0001). Among tumors, methylation class membership was significantly associated with lung tissue asbestos body burden (P < 0.03), and significantly predicted survival (likelihood ratio P < 0.01). Consistent with prior work, asbestos burden was associated with an increased risk of death (hazard ratio, 1.4; 95% confidence interval, 1.1-1.8). Our results have shown that methylation profiles powerfully differentiate diseased pleura from nontumor pleura and that asbestos burden and methylation profiles are independent predictors of mesothelioma patient survival. We have added to the growing body of evidence that cellular epigenetic dysregulation is a critical mode of action for asbestos in the induction of pleural mesothelioma. Importantly, these findings hold great promise for using epigenetic profiling in the diagnosis and prognosis of human cancers.


Assuntos
Amianto/análise , Mesotelioma/genética , Neoplasias Pleurais/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Ilhas de CpG , Metilação de DNA , Epigênese Genética , Feminino , Humanos , Pulmão/química , Masculino , Mesotelioma/química , Mesotelioma/cirurgia , Pessoa de Meia-Idade , Pleura/química , Neoplasias Pleurais/química , Neoplasias Pleurais/cirurgia
20.
Neuro Oncol ; 11(5): 477-87, 2009 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-19139420

RESUMO

Development of model systems that recapitulate the molecular heterogeneity observed among glioblastoma multiforme (GBM) tumors will expedite the testing of targeted molecular therapeutic strategies for GBM treatment. In this study, we profiled DNA copy number and mRNA expression in 21 independent GBM tumor lines maintained as subcutaneous xenografts (GBMX), and compared GBMX molecular signatures to those observed in GBM clinical specimens derived from the Cancer Genome Atlas (TCGA). The predominant copy number signature in both tumor groups was defined by chromosome-7 gain/chromosome-10 loss, a poor-prognosis genetic signature. We also observed, at frequencies similar to that detected in TCGA GBM tumors, genomic amplification and overexpression of known GBM oncogenes, such as EGFR, MDM2, CDK6, and MYCN, and novel genes, including NUP107, SLC35E3, MMP1, MMP13, and DDX1. The transcriptional signature of GBMX tumors, which was stable over multiple subcutaneous passages, was defined by overexpression of genes involved in M phase, DNA replication, and chromosome organization (MRC) and was highly similar to the poor-prognosis mitosis and cell-cycle module (MCM) in GBM. Assessment of gene expression in TCGA-derived GBMs revealed overexpression of MRC cancer genes AURKB, BIRC5, CCNB1, CCNB2, CDC2, CDK2, and FOXM1, which form a transcriptional network important for G2/M progression and/or checkpoint activation. Our study supports propagation of GBM tumors as subcutaneous xenografts as a useful approach for sustaining key molecular characteristics of patient tumors, and highlights therapeutic opportunities conferred by this GBMX tumor panel for testing targeted therapeutic strategies for GBM treatment.


Assuntos
Neoplasias Encefálicas/genética , Dosagem de Genes , Glioblastoma/genética , RNA Mensageiro/análise , Animais , Proliferação de Células , Amplificação de Genes , Humanos , Análise de Sequência com Séries de Oligonucleotídeos , Transcrição Gênica , Transplante Heterólogo
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