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1.
Transl Lung Cancer Res ; 5(5): 511-516, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27826532

RESUMO

Circulating free DNA (cfDNA) is obtained from serum or plasma by non-invasive methods such as a simple blood draw, a technique known as "liquid biopsy". Genetic analyses of driver alterations in cfDNA have proved very effective to predict survival and treatment response of cancer patients according to tumoral cfDNA burden in blood. Non-small cell lung cancer (NSCLC) patients with higher concentration of tumoral cfDNA in blood have, on average, shorter progression-free survival (PFS) and overall survival (OS). Regarding specific genetic alterations, KRAS proto-oncogene, GTPase (KRAS) is one of the main genes involved in NSCLC and several studies have been performed to determine its value as a predictive and prognostic biomarker in liquid biopsy. Unfortunately, to date no strong conclusions can be drawn since they have yielded contradictory results. Therefore, further investigations are necessary to establish the value of KRAS testing in liquid biopsy as prognostic or predictive factor in NSCLC. Herein, we review the current knowledge on the importance of KRAS as prognostic and predictive biomarker using non-invasive approaches and the scientific data available regarding its application in clinical practice for treatment of NSCLC.

2.
Oncotarget ; 7(35): 56619-56627, 2016 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-27447748

RESUMO

Approximately 50% of metastatic melanoma patients harbor BRAF mutations. Several treatment options including the combination of BRAF and MEK inhibitors (BRAF/MEKi) and immunotherapy (mainly anti CTLA-4 and anti PD-1 antibodies), have been shown to improve survival in these patients. Although preclinical data support the synergistic effect of both modalities in combination, data confirming the activity and tolerability of these combinations are not yet available in the clinical setting. Herein, we report the case of a melanoma patient treated with sequential BRAF/MEKi (dabrafenib plus trametinib) followed by the anti CTLA-4 antibody ipilimumab who achieved a pathological complete response. Unfortunately, the patient died due to fatal gastrointestinal (GI) toxicity. Analysis of the BRAFV600E mutation in circulating tumoral DNA (ctDNA) from peripheral blood samples and serial tumor tissue biopsies throughout treatment demonstrated a good correlation with clinical evolution.


Assuntos
Trato Gastrointestinal/efeitos dos fármacos , Imidazóis/efeitos adversos , Ipilimumab/efeitos adversos , Melanoma/tratamento farmacológico , Oximas/efeitos adversos , Piridonas/efeitos adversos , Pirimidinonas/efeitos adversos , Neoplasias Cutâneas/tratamento farmacológico , Antineoplásicos Imunológicos/administração & dosagem , Antineoplásicos Imunológicos/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Biópsia , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/secundário , DNA de Neoplasias/sangue , Evolução Fatal , Humanos , Imidazóis/administração & dosagem , MAP Quinase Quinase Quinases/antagonistas & inibidores , Masculino , Melanoma/metabolismo , Pessoa de Meia-Idade , Mutação , Células Neoplásicas Circulantes/metabolismo , Oximas/administração & dosagem , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Piridonas/administração & dosagem , Pirimidinonas/administração & dosagem , Neoplasias Cutâneas/metabolismo , Resultado do Tratamento
3.
PLoS One ; 9(2): e89518, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24586842

RESUMO

The EURTAC trial demonstrated that the tyrosine kinase inhibitor (TKI) erlotinib was superior to chemotherapy as first-line therapy for advanced non-small cell lung cancers (NSCLC) that harbor EGFR activating mutations in a predominantly Caucasian population. Based on EURTAC and several Asian trials, anti-EGFR TKIs are standard of care for EGFR mutation-positive NSCLC. We sought to validate a rapid multiplex EGFR mutation assay as a companion diagnostic assay to select patients for this therapy. Samples from the EURTAC trial were prospectively screened for EGFR mutations using a combination of laboratory-developed tests (LDTs), and tested retrospectively with the cobas EGFR mutation test (EGFR PCR test). The EGFR PCR test results were compared to the original LDT results and to Sanger sequencing, using a subset of specimens from patients screened for the trial. Residual tissue was available from 487 (47%) of the 1044 patients screened for the trial. The EGFR PCR test showed high concordance with LDT results with a 96.3% overall agreement. The clinical outcome of patients who were EGFR-mutation detected by the EGFR PCR test was very similar to the entire EURTAC cohort. The concordance between the EGFR PCR test and Sanger sequencing was 90.6%. In 78.9% of the discordant samples, the EGFR PCR test result was confirmed by a sensitive deep sequencing assay. This retrospective study demonstrates the clinical utility of the EGFR PCR test in the accurate selection of patients for anti-EGFR TKI therapy. The EGFR PCR test demonstrated improved performance relative to Sanger sequencing.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Carcinoma Pulmonar de Células não Pequenas/genética , Receptores ErbB/genética , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Cloridrato de Erlotinib , Feminino , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Masculino , Pessoa de Meia-Idade , Mutação/genética , Reação em Cadeia da Polimerase/métodos , Estudos Prospectivos , Inibidores de Proteínas Quinases/uso terapêutico , Quinazolinas/uso terapêutico , Estudos Retrospectivos , Análise de Sequência de DNA/métodos
4.
Lung Cancer ; 82(1): 38-43, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23927883

RESUMO

OBJECTIVES: Epidermal growth factor receptor (EGFR) mutations have been identified in lung adenocarcinomas and are associated with high response chance to EGFR tyrosine kinase inhibitors. EGFR mutations can be detected in tumour tissue, cytology specimens and blood from lung cancer patients. Thus far, EGFR mutation analysis has not been systematically demonstrated for sputum samples. The aim of the present study was to determine whether EGFR mutation analysis is attainable on sputum samples, employing different assays in a multicenter study. MATERIALS AND METHODS: Sputum DNA from 10 lung cancer patients with confirmed EGFR mutation in their tumour tissue, 10 lung cancer patients without evidence of an EGFR mutation, and 10 patients with chronic obstructive pulmonary disease (COPD) was used for mutation analysis by Cycleave PCR, COLD-PCR, PangaeaBiotech SL Technology (PST), and High Resolution Melting, respectively. Targeted resequencing (TruSeq Amplicon Cancer Panel) and droplet digital PCR were additionally performed on the 10 samples with EGFR mutation. RESULTS: Dependent on the assay, EGFR mutations could be detected in 30-50% of the sputum samples of patients with EGFR mutations. The different techniques revealed consistent results, with slightly higher sensitivity for PST. Neither the lung cancer patients without EGFR mutation nor the COPD controls tested positive for EGFR mutations in their sputum samples, indicating high clinical specificity of all assays. CONCLUSION: EGFR mutations can be detected in sputum samples from patients with EGFR-mutated non-small cell lung cancer, which may replace biopsy procedure for some patients.


Assuntos
Adenocarcinoma/diagnóstico , Receptores ErbB/genética , Neoplasias Pulmonares/diagnóstico , Neoplasias de Células Escamosas/diagnóstico , Escarro/metabolismo , Adenocarcinoma/genética , Idoso , Estudos de Casos e Controles , DNA/genética , DNA/isolamento & purificação , Análise Mutacional de DNA , Humanos , Neoplasias Pulmonares/genética , Pessoa de Meia-Idade , Técnicas de Diagnóstico Molecular , Neoplasias de Células Escamosas/genética , Reação em Cadeia da Polimerase , Doença Pulmonar Obstrutiva Crônica/genética , Sensibilidade e Especificidade , Método Simples-Cego , Temperatura de Transição
5.
J Thorac Oncol ; 8(3): 295-300, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23407556

RESUMO

INTRODUCTION: Lung adenocarcinoma patients harboring EGFR activating mutations attain improved progression-free survival (PFS) with treatment with epidermal growth factor receptor tyrosine kinase inhibitors. However, patients ultimately relapse, indicating that other genetic factors could influence outcome in such patients. We hypothesized that PFS could be influenced by the expression of genes in DNA repair pathways. METHODS: We examined the mRNA expression of C terminus-binding protein-interacting protein and Lin11, Isl-1, and Mec-3 domain only 4 (LMO4) in pretreatment tumor samples from 91 erlotinib-treated advanced non-small-cell lung cancer patients with EGFR mutations in whom breast cancer gene 1 (BRCA1) expression and the concomitant presence of the EGFR T790M mutation had previously been assessed. Gene expression was analyzed by polymerase chain reaction, using ß-actin as endogenous gene. Results were correlated with PFS and overall survival. RESULTS: In patients with low LMO4 levels, PFS was 13 months, whereas it was not reached for those with high LMO4 levels (p = 0.03). In patients with low levels of both BRCA1 and LMO4, PFS was 19 months whereas it was not reached in those with low BRCA1 and high LMO4 mRNA levels (p = 0.04). In patients with high BRCA1 and low LMO4 levels, PFS was 8 months, whereas it was 18 months in those with high levels of both genes (p = 0.03). CONCLUSIONS: Low BRCA1 and high LMO4 levels were associated with longer PFS to erlotinib. Baseline assessment of BRCA1 and LMO4 mRNA expression can help predict outcome to erlotinib.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteína BRCA1/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Proteínas de Transporte/genética , Receptores ErbB/genética , Proteínas com Domínio LIM/genética , Mutação/genética , Proteínas Nucleares/genética , Quinazolinas/uso terapêutico , RNA Mensageiro/genética , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/genética , Adenocarcinoma/mortalidade , Adenocarcinoma Bronquioloalveolar/tratamento farmacológico , Adenocarcinoma Bronquioloalveolar/genética , Adenocarcinoma Bronquioloalveolar/mortalidade , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Carcinoma de Células Grandes/tratamento farmacológico , Carcinoma de Células Grandes/genética , Carcinoma de Células Grandes/mortalidade , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Endodesoxirribonucleases , Cloridrato de Erlotinib , Feminino , Seguimentos , Perfilação da Expressão Gênica , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/mortalidade , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Análise de Sequência com Séries de Oligonucleotídeos , Prognóstico , Estudos Prospectivos , Inibidores de Proteínas Quinases/uso terapêutico , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Taxa de Sobrevida , Adulto Jovem
6.
Clin Lung Cancer ; 14(3): 205-14, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23122493

RESUMO

Epidermal growth factor receptor (EGFR) gene mutations and increased EGFR copy numbers have been associated with a favorable response to EGFR tyrosine kinase inhibitors (TKI) in patients with non-small-cell lung cancer (NSCLC), and several markers have been identified that predict response to treatment. Lung adenocarcinomas also harbor activating mutations in the downstream GTPase, v-Ki-ras2 Kirsten rat sarcoma viral oncogene (KRAS), and mutations in EGFR and KRAS appear to be mutually exclusive. Even though KRAS mutations were identified in NSCLC tumors more than 20 years ago, we have only just begun to appreciate the clinical value of determining KRAS tumor status. Recent studies indicate that patients with mutant KRAS tumors fail to benefit from adjuvant chemotherapy and do not respond to EGFR inhibitors. There is a clear need for therapies specifically developed for patients with KRAS-mutant NSCLC. In this review, we summarize the clinical and pathologic characteristics of patients with NSCLC and with KRAS mutations, describe work that explores the predictive and prognostic influence of KRAS mutations, and provide an overview of the "synthetic lethal" interactions and current approaches to targeting KRAS-mutant NSCLC.


Assuntos
Adenocarcinoma/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Pulmonares/genética , Mutação , Proteínas Proto-Oncogênicas/genética , Proteínas ras/genética , Animais , Humanos , Proteínas Proto-Oncogênicas p21(ras)
7.
Transl Lung Cancer Res ; 2(2): 65-71, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25806217

RESUMO

Circulating tumor cells (CTCs) can be detected in the blood of many cancer patients and play a key role in metastasis. In addition, after the development of technologies with the necessary sensitivity and reproducibility, the diagnostic potential of these cells is being actively explored. Recently, the U.S. Food and Drug Administration has approved the CellSearch(®) System, based on magnetic beads coated with epithelial cell-adhesion molecule (EpCAM) antibody. Despite its usefulness, this system can miss CTCs that lose epithelial antigens due to the epithelial-mesenchymal transition and, in the case of advanced NSCLC, CTCs positivity can be demonstrated only in 30-50% of patients. In an effort to overcome these drawbacks, new methods are being developed. In this study, we have evaluated CK-coated beads as a system to isolate CTCs from lung cancer patients in the clinical setting, and have evaluated if they can be a useful source of material for genetic testing. We were able to identify CTCs in 17 of the 30 patients included in the study (57%), with a range of 1 to 7 cells. In two of them, we found only CTCs with an EMT pattern. CTC positivity seemed to correlate with the clinical history of the malignancy. CTCs could be detected in more than 80% of stage III-IV lung cancer patients at presentation or in blood samples taken immediately after surgery. The percentage dropped to 13% in patients responding to chemotherapy or TKIs, raising again to 57% after tumor progression. Finally, we tested the CTCs isolated from 8 patients for EGFR and k-ras mutations, but gene amplification was successful only in the 3 patients with 4 or more CTCs.

8.
Pharmacogenomics ; 13(7): 789-802, 2012 May.
Artigo em Inglês | MEDLINE | ID: mdl-22594511

RESUMO

Lung cancer is a lethal disease, and most cases have already disseminated at the time of diagnosis. Driver mutations in the EGFR tyrosine kinase domain (mainly deletions in exon 19 and L858R mutation in exon 21) have been identified in lung adenocarcinomas, mostly in never smokers, at frequencies of 20-60%. The EGFR tyrosine kinase inhibitors (TKIs) gefitinib or erlotinib attain a response rate of 70% and progression-free survival of 9-13 months, although there are subgroups of patients with long-lasting remissions. No significant correlation between EGFR overexpression and response to treatment has been found, while controversial results have been reported regarding EGFR gene amplification. The pretreatment presence of the T790M mutation, initially identified as an acquired resistance mutation to treatment with EGFR TKIs, has also been reported and may indicate a genetically distinct disease. Finally, other genetic factors, such as mRNA expression of BRCA1 and components of the NF-κB pathway, can modulate response to EGFR TKIs in EGFR-mutated patients.


Assuntos
Carcinoma Pulmonar de Células não Pequenas , Receptores ErbB , Neoplasias Pulmonares , Inibidores de Proteínas Quinases , Biomarcadores Farmacológicos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/genética , Cloridrato de Erlotinib , Gefitinibe , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Farmacogenética , Inibidores de Proteínas Quinases/uso terapêutico , Quinazolinas/uso terapêutico
9.
J Transl Med ; 9: 163, 2011 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-21951562

RESUMO

BACKGROUND: Metastatic non-small-cell lung cancer (NSCLC) has a dismal prognosis. EGFR is overexpressed or mutated in a large proportion of cases. Downstream components of the EGFR pathway and crosstalk with the NF-κB pathway have not been examined at the clinical level. We explored the prognostic significance of the mRNA expression of nine genes in the EGFR and NF-κB pathways and of BRCA1 and RAP80 in patients in whom EGFR and K-ras gene status had previously been determined. In addition, NFKBIA and DUSP22 gene status was also determined. METHODS: mRNA expression of the eleven genes was determined by QPCR in 60 metastatic NSCLC patients and in nine lung cancer cell lines. Exon 3 of NFKBIA and exon 6 of DUSP22 were analyzed by direct sequencing. Results were correlated with outcome to platinum-based chemotherapy in patients with wild-type EGFR and to erlotinib in those with EGFR mutations. RESULTS: BRCA1 mRNA expression was correlated with EZH2, AEG-1, Musashi-2, CYLD and TRAF6 expression. In patients with low levels of both BRCA1 and AEG-1, PFS was 13.02 months, compared to 5.4 months in those with high levels of both genes and 7.7 months for those with other combinations (P=0.025). The multivariate analysis for PFS confirmed the prognostic role of high BRCA1/AEG-1 expression (HR, 3.1; P=0.01). Neither NFKBIA nor DUSP22 mutations were found in any of the tumour samples or cell lines. CONCLUSIONS: The present study provides a better understanding of the behaviour of metastatic NSCLC and identifies the combination of BRCA1 and AEG-1 expression as a potential prognostic model.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Receptores ErbB/metabolismo , Regulação Neoplásica da Expressão Gênica , Genes Neoplásicos/genética , Neoplasias Pulmonares/genética , NF-kappa B/metabolismo , Transdução de Sinais/genética , Adulto , Idoso , Proteína BRCA1/genética , Proteína BRCA1/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Intervalo Livre de Doença , Receptores ErbB/genética , Feminino , Humanos , Neoplasias Pulmonares/patologia , Masculino , Proteínas de Membrana , Pessoa de Meia-Idade , Análise Multivariada , Mutação/genética , NF-kappa B/genética , Metástase Neoplásica , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA
10.
Clin Cancer Res ; 17(5): 1160-8, 2011 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-21233402

RESUMO

PURPOSE: Advanced non-small-cell lung cancer (NSCLC) patients harboring epidermal growth factor receptor (EGFR) mutations (deletion in exon 19 or L858R) show an impressive progression-free survival of 14 months when treated with erlotinib. However, the presence of EGFR mutations can only imperfectly predict outcome. We hypothesized that progression-free survival could be influenced both by the pretreatment EGFR T790M mutation and by components of DNA repair pathways. EXPERIMENTAL DESIGN: We assessed the T790M mutation in pretreatment diagnostic specimens from 129 erlotinib-treated advanced NSCLC patients with EGFR mutations. The expression of eight genes and two proteins involved in DNA repair and four receptor tyrosine kinases was also examined. RESULTS: The EGFR T790M mutation was observed in 45 of 129 patients (35%). Progression-free survival was 12 months in patients with and 18 months in patients without the T790M mutation (P = 0.05). Progression-free survival was 27 months in patients with low BRCA1 mRNA levels, 18 months in those with intermediate levels, and 10 months in those with high levels (P = 0.02). In the multivariate analysis, the presence of the T790M mutation (HR, 4.35; P = 0.001), intermediate BRCA1 levels (HR, 8.19; P < 0.0001), and high BRCA1 levels (HR, 8.46; P < 0.0001) emerged as markers of shorter progression-free survival. CONCLUSIONS: Low BRCA1 levels neutralized the negative effect of the T790M mutation and were associated with longer progression-free survival to erlotinib. We advocate baseline assessment of the T790M mutation and BRCA1 expression to predict outcome and provide alternative individualized treatment to patients based on T790M mutations and BRCA1 expression.


Assuntos
Proteína BRCA1/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Receptores ErbB/genética , Genes BRCA1 , Neoplasias Pulmonares/genética , Mutação , Quinazolinas/uso terapêutico , Adulto , Idoso , Idoso de 80 Anos ou mais , Antineoplásicos/uso terapêutico , Biomarcadores Tumorais , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Reparo do DNA/genética , Intervalo Livre de Doença , Cloridrato de Erlotinib , Feminino , Expressão Gênica , Genes erbB-1 , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Masculino , Pessoa de Meia-Idade , Inibidores de Proteínas Quinases/uso terapêutico , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Resultado do Tratamento
11.
J Transl Med ; 8: 135, 2010 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-21167064

RESUMO

BACKGROUND: Immunohistochemistry (IHC) with mutation-specific antibodies may be an ancillary method of detecting EGFR mutations in lung cancer patients. METHODS: EGFR mutation status was analyzed by DNA assays, and compared with IHC results in five non-small-cell lung cancer (NSCLC) cell lines and tumor samples from 78 stage IV NSCLC patients. RESULTS: IHC correctly identified del 19 in the H1650 and PC9 cell lines, L858R in H1975, and wild-type EGFR in H460 and A549, as well as wild-type EGFR in tumor samples from 22 patients. IHC with the mAb against EGFR with del 19 was highly positive for the protein in all 17 patients with a 15-bp (ELREA) deletion in exon 19, whereas in patients with other deletions, IHC was weakly positive in 3 cases and negative in 9 cases. IHC with the mAb against the L858R mutation showed high positivity for the protein in 25/27 (93%) patients with exon 21 EGFR mutations (all with L858R) but did not identify the L861Q mutation in the remaining two patients. CONCLUSIONS: IHC with mutation-specific mAbs against EGFR is a promising method for detecting EGFR mutations in NSCLC patients. However these mAbs should be validated with additional studies to clarify their possible role in routine clinical practice for screening EGFR mutations in NSCLC patients.


Assuntos
Anticorpos Antineoplásicos/imunologia , Especificidade de Anticorpos/imunologia , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/imunologia , Receptores ErbB/genética , Neoplasias Pulmonares/genética , Mutação/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Monoclonais , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Análise Mutacional de DNA , Éxons/genética , Feminino , Humanos , Imuno-Histoquímica , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Deleção de Sequência
12.
Discov Med ; 8(43): 181-4, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-20040267

RESUMO

Certain mutations in the epidermal growth factor receptor (EGFR) gene confer hypersensitivity to the tyrosine kinase inhibitors gefitinib and erlotinib in patients with advanced non-small cell lung cancer. Large-scale screening for EGFR mutations in such patients is feasible for predicting response to TKIs and thus guiding treatment.


Assuntos
Receptores ErbB/genética , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Mutação/genética , Receptores ErbB/sangue , Humanos , Reação em Cadeia da Polimerase
13.
N Engl J Med ; 361(10): 958-67, 2009 Sep 03.
Artigo em Inglês | MEDLINE | ID: mdl-19692684

RESUMO

BACKGROUND: Activating mutations in the epidermal growth factor receptor gene (EGFR) confer hypersensitivity to the tyrosine kinase inhibitors gefitinib and erlotinib in patients with advanced non-small-cell lung cancer. We evaluated the feasibility of large-scale screening for EGFR mutations in such patients and analyzed the association between the mutations and the outcome of erlotinib treatment. METHODS: From April 2005 through November 2008, lung cancers from 2105 patients in 129 institutions in Spain were screened for EGFR mutations. The analysis was performed in a central laboratory. Patients with tumors carrying EGFR mutations were eligible for erlotinib treatment. RESULTS: EGFR mutations were found in 350 of 2105 patients (16.6%). Mutations were more frequent in women (69.7%), in patients who had never smoked (66.6%), and in those with adenocarcinomas (80.9%) (P<0.001 for all comparisons). The mutations were deletions in exon 19 (62.2%) and L858R (37.8%). Median progression-free survival and overall survival for 217 patients who received erlotinib were 14 months and 27 months, respectively. The adjusted hazard ratios for the duration of progression-free survival were 2.94 for men (P<0.001); 1.92 for the presence of the L858R mutation, as compared with a deletion in exon 19 (P=0.02); and 1.68 for the presence of the L858R mutation in paired serum DNA, as compared with the absence of the mutation (P=0.02). The most common adverse events were mild rashes and diarrhea; grade 3 cutaneous toxic effects were recorded in 16 patients (7.4%) and grade 3 diarrhea in 8 patients (3.7%). CONCLUSIONS: Large-scale screening of patients with lung cancer for EGFR mutations is feasible and can have a role in decisions about treatment.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Receptores ErbB/genética , Neoplasias Pulmonares/genética , Mutação , Inibidores de Proteínas Quinases/uso terapêutico , Quinazolinas/uso terapêutico , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Diarreia/induzido quimicamente , Receptores ErbB/antagonistas & inibidores , Cloridrato de Erlotinib , Exantema/induzido quimicamente , Feminino , Humanos , Estimativa de Kaplan-Meier , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/mortalidade , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Estudos Prospectivos , Inibidores de Proteínas Quinases/efeitos adversos , Quinazolinas/efeitos adversos , Distribuição por Sexo , Análise de Sobrevida , Adulto Jovem
14.
Cancer Lett ; 281(2): 183-7, 2009 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-19303206

RESUMO

HT-29 M6 colon cancer cells differentiate to a mucus-secreting phenotype in culture. We found that the pattern of cyclin D1 expression in HT-29 M6 cells did not correlate with instances of cell proliferation but was specifically induced during a dedifferentiation process following disaggregation of epithelial cell layers, even under conditions that did not allow cell cycle reentrance. Interestingly, ectopic expression of cyclin D1 in differentiated cells led to the inhibition of the transcriptional activity of differentiation gene promoters, such as the mucin MUC1. We thus propose that the overexpression of cyclin D1 found in colon cancer favours tumour dedifferentiation as one mechanism of tumour progression.


Assuntos
Desdiferenciação Celular/genética , Neoplasias do Colo/genética , Ciclina D1/genética , Regulação Neoplásica da Expressão Gênica/fisiologia , Muco/metabolismo , Western Blotting , Ciclo Celular/fisiologia , Diferenciação Celular/genética , Linhagem Celular Tumoral , Proliferação de Células , Neoplasias do Colo/metabolismo , Ciclina D1/metabolismo , Progressão da Doença , Citometria de Fluxo , Expressão Gênica , Humanos , Mucina-1/genética , Regiões Promotoras Genéticas , Transfecção
15.
J Thorac Oncol ; 3(11): 1224-35, 2008 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-18978556

RESUMO

BACKGROUND: Detection of epidermal growth factor receptor (EGFR) mutations in advanced non-small cell lung cancer (NSCLC) patients has relied on DNA purification from biopsies, amplification, and sequencing. However, the number of tumor cells in a sample is often insufficient for EGFR assessment. METHODS: We prospectively screened 1380 NSCLC patients for EGFR mutations but found that 268 were not evaluable because of insufficient tumor tissue. We therefore developed and validated a method of detecting EGFR mutations in these samples. Tumor cells were microdissected into polymerase chain reaction buffer and amplified. EGFR mutations were detected by length analysis of fluorescently labeled polymerase chain reaction products and TaqMan assay. RESULTS: We determined EGFR status in 217 (81%) of the 268 primary NSCLC samples not evaluable in our original study-fresh and paraffin-embedded with less than 150 cells. Exon 19 deletions were detected in 11.5% of patients and exon 21 L858R mutations in 5.5%. In addition, the exon 20 T790M mutation was detected in 6 of 15 (40%) patients at the time of progression to erlotinib. The primary, sensitive mutation was present in all tumor cells, whereas the T790M mutation was absent in some groups. CONCLUSIONS: The method presented here eliminates the need for DNA purification and allows for detection of EGFR mutations in samples containing as few as eight cancer cells.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Receptores ErbB/genética , Neoplasias Pulmonares/genética , Mutação/genética , Reação em Cadeia da Polimerase/métodos , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/genética , Adenocarcinoma/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/patologia , DNA de Neoplasias/genética , Receptores ErbB/antagonistas & inibidores , Cloridrato de Erlotinib , Éxons , Feminino , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Estudos Prospectivos , Inibidores de Proteínas Quinases/uso terapêutico , Quinazolinas/uso terapêutico , Sensibilidade e Especificidade , Taxa de Sobrevida
16.
J Cell Physiol ; 212(1): 42-50, 2007 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-17311291

RESUMO

Tumor cell dedifferentiation-such as the loss of cell-to-cell adhesion in epithelial tumors-is associated with tumor progression. To better understand the mechanisms that maintain carcinoma cells in a differentiated state, we have dissected in vitro differentiation pathways in the mucus-secretor HT-29 M6 colon cancer cell line, which spontaneously differentiates in postconfluent cultures. By lowering the extracellular calcium concentration to levels that prevent intercellular adhesion and epithelial polarization, our results reveal that differentiation is calcium-dependent and involves: (i) a process of cell cycle exit to G(0) and (ii) the induction of a transcriptional program of differentiation gene expression (i.e., mucins MUC1 and MUC5AC, and the apical membrane peptidase DPPIV). In calcium-deprived, non-differentiated postconfluent cultures, differentiation gene promoters are repressed by a trichostatin A (TSA)-sensitive mechanism, indicating that loss of gene expression by dedifferentiation is driven by histone deacetylases (HDAC). Since TSA treatment or extracellular calcium restoration allow gene promoter activation to similar levels, we suggest that induction of differentiation is one mechanism of HDAC inhibitor antitumor action. Moreover, transcriptional de-repression can also be induced in non-differentiating culture conditions by overexpressing the cyclin-dependent kinase inhibitor p27(KIP1), which is normally induced during spontaneous differentiation. Since p27(KIP1) downregulation in colon cancer is associated with poor prognosis independently of tumor cell division rates, we propose that p27 (KIP1) may prevent tumor progression by, at least in part, enhancing the expression of some differentiation genes. Therefore, the HT-29 M6 model allows the identification of some basic mechanisms of cancer cell differentiation control, so far revealing HDAC and p27(KIP1) as key regulatory factors of differentiation gene expression.


Assuntos
Neoplasias do Colo/metabolismo , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Ácidos Hidroxâmicos/farmacologia , Muco/metabolismo , Transcrição Gênica/fisiologia , Cálcio/metabolismo , Ciclo Celular , Diferenciação Celular , Regulação Neoplásica da Expressão Gênica , Células HT29 , Histona Desacetilases/metabolismo , Humanos , Intestinos/citologia , Regiões Promotoras Genéticas/fisiologia
17.
Int J Cancer ; 119(7): 1607-15, 2006 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-16671091

RESUMO

Genetic instability leads to tumor heterogeneity, which in turn provides a source of cell variants responsible for drug resistance. However, the source of resistant cells during the process of acquired resistance is poorly understood. Our aim has been to characterize the mechanism by which acquired resistance to methotrexate emerges during the course of cancer cell treatment in vitro. We recently demonstrated that, in vitro, HT-29 colon cancer cells become transiently sensitive to methotrexate by depleting the extracellular milieu of survival factors; on the other hand, the cell population under treatment can reversibly adapt to grow below a critical cell density in the presence of the drug. Here, we show that this adapted cell population gives rise to permanent resistant populations through repeated cycles of cell death and growth. This increased cell turnover, but not merely cell proliferation, is required for the appearance of increasing degrees of stable resistance that are progressively selected by drug pressure. Such a process, taking place in multiple steps, is here designated "dynamic selection." The analysis of sensitive and resistant HT-29 cell populations revealed that methotrexate induces genomic instability--characterized by centrosome amplification and aberrant chromosome recombination--leading to a low-level amplification of the 5q chromosome arm as one of the earliest genetic events selected during treatment. Therefore, this model provides a mechanism by which a tumor cell population lacking resistant subpopulations before treatment is able to acquire the genetic changes required for stable drug resistance.


Assuntos
Resistencia a Medicamentos Antineoplásicos , Variação Genética/genética , Metotrexato/farmacologia , Neoplasias/genética , Neoplasias/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Instabilidade Cromossômica/genética , Dipiridamol/farmacologia , Humanos , Neoplasias/classificação , Nucleosídeos/farmacologia , Seleção Genética
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