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1.
Cell Rep ; 30(13): 4551-4566.e7, 2020 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-32234487

RESUMO

Loss-of-function mutations in the SDHB subunit of succinate dehydrogenase predispose patients to aggressive tumors characterized by pseudohypoxic and hypermethylator phenotypes. The mechanisms leading to DNA hypermethylation and its contribution to SDH-deficient cancers remain undemonstrated. We examine the genome-wide distribution of 5-methylcytosine and 5-hydroxymethylcytosine and their correlation with RNA expression in SDHB-deficient tumors and murine Sdhb-/- cells. We report that DNA hypermethylation results from TET inhibition. Although it preferentially affects PRC2 targets and known developmental genes, PRC2 activity does not contribute to the DNA hypermethylator phenotype. We also prove, in vitro and in vivo, that TET silencing, although recapitulating the methylation profile of Sdhb-/- cells, is not sufficient to drive their EMT-like phenotype, which requires additional HIF2α activation. Altogether, our findings reveal synergistic roles of TET repression and pseudohypoxia in the acquisition of metastatic traits, providing a rationale for targeting HIF2α and DNA methylation in SDH-associated malignancies.

2.
J Clin Endocrinol Metab ; 105(3)2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31678991

RESUMO

CONTEXT: Pheochromocytomas and paragangliomas (PPGLs) are neuroendocrine tumors explained by germline or somatic mutations in about 70% of cases. Patients with SDHB mutations are at high risk of developing metastatic disease, yet no reliable tumor biomarkers are available to predict tumor aggressiveness. OBJECTIVE: We aimed at identifying long noncoding RNAs (lncRNAs) specific for PPGL molecular groups and metastatic progression. DESIGN AND METHODS: To analyze the expression of lncRNAs, we used a mining approach of transcriptome data from a well-characterized series of 187 tumor tissues. Clustering consensus analysis was performed to determine a lncRNA-based classification, and informative transcripts were validated in an independent series of 51 PPGLs. The expression of metastasis-related lncRNAs was confirmed by RT-qPCR. Receiver operating characteristic (ROC) curve analysis was used to estimate the predictive accuracy of potential markers. MAIN OUTCOME MEASURE: Univariate/multivariate and metastasis-free survival (MFS) analyses were carried out for the assessment of risk factors and clinical outcomes. RESULTS: Four lncRNA-based subtypes strongly correlated with mRNA expression clusters (chi-square P-values from 1.38 × 10-32 to 1.07 × 10-67). We identified one putative lncRNA (GenBank: BC063866) that accurately discriminates metastatic from benign tumors in patients with SDHx mutations (area under the curve 0.95; P = 4.59 × 10-05). Moreover, this transcript appeared as an independent risk factor associated with poor clinical outcome of SDHx carriers (log-rank test P = 2.29 × 10-05). CONCLUSION: Our findings extend the spectrum of transcriptional dysregulations in PPGL to lncRNAs and provide a novel biomarker that could be useful to identify potentially metastatic tumors in patients carrying SDHx mutations.

3.
Hepatology ; 2019 Dec 25.
Artigo em Inglês | MEDLINE | ID: mdl-31875970

RESUMO

BACKGROUND & AIMS: Intrahepatic cholangiocarcinoma (ICC) is asevere malignant tumor in which the standard therapies are mostly ineffective. The biological significance of the desmoplastic tumor microenvironment (TME) of ICC has been stressed, but was insufficiently taken into account in the search for classifications of ICC adapted to clinical trial design. We investigated the heterogeneous tumor stroma composition and built a TME-based classification of ICC tumors, which detects potentially targetable ICC subtypes. METHODS: We established the bulk gene expression profiles of 78 ICCs. Epithelial and stromal compartments of 23 ICCs were laser microdissected. We quantified 14 gene-expression signatures of the TME and those of 3 functional indicators (liver activity, inflammation, immune resistance). The cell population abundances were quantified using the Microenvironment Cell Populations (MCP)-counter package and compared with immunohistochemistry. We performed an unsupervised TME-based classification of 198 ICCs (training set) and 368 ICCs (validation set). We determined immune response and signaling features of the different immune subtypes by functional annotations. RESULTS: We showed that a set of 198 ICCs could be classified into 4 TME-based subtypes related to distinct immune escape mechanisms and patient outcomes. The validity of these immune subtypes was confirmed over an independent set of 368 ICCs and by immunohistochemical analysis of 64 ICC tissue samples. About 45% of ICCs displayed an immune desert phenotype. The other subtypes differed in the nature (lymphoid, myeloid, mesenchymal) and abundance of tumor infiltrating cells. The inflamed subtype (11%) presented a massive T-lymphocyte infiltration, an activation of inflammatory and immune checkpoint pathways, and was associated with the longest patient survival. CONCLUSION: We revealed the existence of an inflamed ICC subtype, which is potentially treatable with checkpoint blockade immunotherapy.

4.
Cancers (Basel) ; 11(6)2019 Jun 08.
Artigo em Inglês | MEDLINE | ID: mdl-31181806

RESUMO

The management of locally advanced head and neck squamous cell carcinoma (HNSCC) with Cetuximab, a monoclonal antibody targeting the epidermal growth factor receptor (EGFR), achieves only moderate response rates, and clinical trials that evaluated EGFR-blockade with tyrosine kinase inhibitors (TKI) yielded disappointing results. Inter-tumor heterogeneity may hinder the therapeutic efficiency of anti-EGFR treatments. HNSCC heterogeneity was addressed in several studies, which all converged towards the definition of molecular subgroups. They include the basal subgroup, defined by the deregulated expression of factors involved in the EGFR signaling pathway, including the epiregulin EGFR ligand encoded by the EREG gene. These observations indicate that basal tumors could be more sensitive to anti-EGFR treatments. To test this hypothesis, we performed a screen of a representative collection of basal versus non-basal HNSCC cell lines for their sensitivity to several anti-EGFR drugs (Cetuximab, Afatinib, and Gefitinib), tested as monotherapy or in combination with drugs that target closely-linked pathways [Mitogen-activated protein kinase kinase/ extracellular signal-regulated kinases (MEK), mammalian Target of Rapamycine (mTOR) or Human Epidermal growth factor Receptor 2 (HER2)]. Basal-like cell lines were found to be more sensitive to EGFR blockade alone or in combination with treatments that target MEK, mTOR, or HER2. Strikingly, the basal-like status was found to be a better predictor of cell response to EGFR blockade than clinically relevant mutations [e.g., cyclin-dependent kinase Inhibitor 2A (CDKN2A)]. Interestingly, we show that EGFR blockade inhibits EREG expression, and that EREG knock-down decreases basal cell clonogenic survival, suggesting that EREG expression could be a predictive functional marker of sensitivity to EGFR blockade in basal-like HNSCC.

5.
Clin Genitourin Cancer ; 17(5): e981-e994, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31229459

RESUMO

INTRODUCTION: Recent trials have suggested predictive biomarkers in advanced clear-cell renal cell carcinoma (accRCC): International Metastatic RCC Database Consortium (IMDC) good risk or angiogenic gene signature for sunitinib and IMDC intermediate/poor risk for ipilimumab-nivolumab and T-effector cell signature or sarcomatoid dedifferentiation for atezolizumab-bevacizumab. We hypothesized that earlier described molecular subtypes, ccrcc1 to ccrcc4, could provide similar information as a single generic biomarker and molecularly characterize the heterogeneous intermediate-risk group. PATIENTS AND METHODS: Patients with accRCC treated with systemic therapies were included. We assessed associations between the 5 biomarkers and their impact on progression-free survival (PFS) and response rate (RR) on first-line sunitinib or pazopanib. The cutoff percentage of sarcomatoid dedifferentiation with optimal discriminative value was determined. RESULTS: In total, 430 patients were included (163 with molecular data). The molecular ccrcc2 subtype identified tumors with higher angiogenic gene expression across IMDC risk groups: prevalence was high in IMDC good risk and low in IMDC poor risk (P < .001). Molecular subtype, IMDC, and angiogenic gene expression had comparable C-indices to predict PFS and RR (range, 60%-66%). The ccrcc2 subtype and angiogenic gene expression were positive predictors of PFS in IMDC intermediate-risk patients (P = .006; P = .04). Immune signature did not differ between IMDC groups, but was strongly correlated with molecular subtype (P = .8 and P = .0007). A cutoff value of 25% sarcomatoid differentiation discriminated tumors with distinct molecular characteristics and therapeutic sensitivity. CONCLUSION: In accRCC, molecular subtypes can explain differences in IMDC risk group, expression of angiogenesis and immune response genes, and sarcomatoid dedifferentiation. They can identify molecularly different patient populations within the heterogeneous IMDC intermediate group and select patients for systemic therapies.


Assuntos
Inibidores da Angiogênese/uso terapêutico , Biomarcadores Tumorais/genética , Carcinoma de Células Renais/tratamento farmacológico , Neoplasias Renais/tratamento farmacológico , Anticorpos Monoclonais Humanizados/uso terapêutico , Bevacizumab/uso terapêutico , Carcinoma de Células Renais/genética , Feminino , Humanos , Neoplasias Renais/genética , Masculino , Terapia de Alvo Molecular , Medicina de Precisão , Pirimidinas/uso terapêutico , Sulfonamidas/uso terapêutico , Análise de Sobrevida , Resultado do Tratamento
6.
Clin Cancer Res ; 25(2): 760-770, 2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30301828

RESUMO

PURPOSE: Pheochromocytomas and paragangliomas (PPGLs) are rare neuroendocrine tumors. Whereas most PPGLs are benign, up to 20% may become metastatic with SDHB- and FH-mutated tumors showing the higher risk. We aimed at determining the contribution of immortalization mechanisms to metastatic progression.Experimental Design: Immortalization mechanisms were investigated in 200 tumors. To identify telomerase (+) tumors, we analyzed genomic alterations leading to transcriptional activation of TERT comprising promoter mutations, hypermethylation and gain copy number. To identify tumors that activated the alternative lengthening of telomere (ALT) mechanism, we combined analyses of telomere length by slot blot, telomere heterogeneity by telomere FISH, and ATRX mutations by next-generation sequencing. Univariate/multivariate and metastasis-free survival (MFS) and overall survival (OS) analyses were carried out for assessment of risk factors and clinical outcomes. RESULTS: Only 37 of 200 (18.5%) tumors achieved immortalization. Telomerase activation occurred in 12 metastatic tumors and was prevalent in SDHB-mutated paragangliomas (P = 2.42e-09). ALT features were present in 25 tumors, mostly pheochromocytomas, regardless of metastatic status or molecular group (P = 0.169), yet ATRX mutations were found preferentially in SDHB/FH-mutated metastatic tumors (P = 0.0014). Telomerase activation and ATRX mutations were independent factors of poor prognosis: MFS (hazard ratio, 48.2 and 33.1; P = 6.50E-07 and 1.90E-07, respectively); OS (hazard ratio, 97.4 and 44.1; P = 4.30E-03 and 2.00E-03, respectively) and were associated with worse MFS and OS (log-rank tests P < 0.0001). CONCLUSIONS: Assessment of telomerase activation and ATRX mutations could be used to identify metastatic PPGLs, particularly in tumors at high risk of progression.


Assuntos
Paraganglioma/genética , Paraganglioma/metabolismo , Feocromocitoma/genética , Feocromocitoma/metabolismo , Telomerase/metabolismo , Proteína Nuclear Ligada ao X/genética , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Metilação de DNA , Análise Mutacional de DNA , Ativação Enzimática , Humanos , Mutação , Estadiamento de Neoplasias , Paraganglioma/mortalidade , Paraganglioma/patologia , Feocromocitoma/mortalidade , Feocromocitoma/patologia , Prognóstico , Regiões Promotoras Genéticas , Sequenciamento Completo do Genoma , Proteína Nuclear Ligada ao X/metabolismo
7.
Cell Mol Gastroenterol Hepatol ; 6(3): 277-300, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30116770

RESUMO

Background & Aims: Recent studies have shown that cancers arise as a result of the positive selection of driver somatic events in tumor DNA, with negative selection playing only a minor role, if any. However, these investigations were concerned with alterations at nonrepetitive sequences and did not take into account mutations in repetitive sequences that have very high pathophysiological relevance in the tumors showing microsatellite instability (MSI) resulting from mismatch repair deficiency investigated in the present study. Methods: We performed whole-exome sequencing of 47 MSI colorectal cancers (CRCs) and confirmed results in an independent cohort of 53 MSI CRCs. We used a probabilistic model of mutational events within microsatellites, while adapting pre-existing models to analyze nonrepetitive DNA sequences. Negatively selected coding alterations in MSI CRCs were investigated for their functional and clinical impact in CRC cell lines and in a third cohort of 164 MSI CRC patients. Results: Both positive and negative selection of somatic mutations in DNA repeats was observed, leading us to identify the expected true driver genes associated with the MSI-driven tumorigenic process. Several coding negatively selected MSI-related mutational events (n = 5) were shown to have deleterious effects on tumor cells. In the tumors in which deleterious MSI mutations were observed despite the negative selection, they were associated with worse survival in MSI CRC patients (hazard ratio, 3; 95% CI, 1.1-7.9; P = .03), suggesting their anticancer impact should be offset by other as yet unknown oncogenic processes that contribute to a poor prognosis. Conclusions: The present results identify the positive and negative driver somatic mutations acting in MSI-driven tumorigenesis, suggesting that genomic instability in MSI CRC plays a dual role in achieving tumor cell transformation. Exome sequencing data have been deposited in the European genome-phenome archive (accession: EGAS00001002477).


Assuntos
Carcinogênese/genética , Neoplasias Colorretais/genética , Instabilidade de Microssatélites , Mutação/genética , Sequências Repetitivas de Ácido Nucleico/genética , Animais , Sequência de Bases , Linhagem Celular Tumoral , Estudos de Coortes , Feminino , Xenoenxertos , Humanos , Masculino , Camundongos , Camundongos Nus , Modelos Estatísticos , Sequenciamento Completo do Exoma
8.
Blood ; 132(5): 469-483, 2018 08 02.
Artigo em Inglês | MEDLINE | ID: mdl-29891534

RESUMO

Chuvash polycythemia is an autosomal recessive form of erythrocytosis associated with a homozygous p.Arg200Trp mutation in the von Hippel-Lindau (VHL) gene. Since this discovery, additional VHL mutations have been identified in patients with congenital erythrocytosis, in a homozygous or compound-heterozygous state. VHL is a major tumor suppressor gene, mutations in which were first described in patients presenting with VHL disease, which is characterized by the development of highly vascularized tumors. Here, we identify a new VHL cryptic exon (termed E1') deep in intron 1 that is naturally expressed in many tissues. More importantly, we identify mutations in E1' in 7 families with erythrocytosis (1 homozygous case and 6 compound-heterozygous cases with a mutation in E1' in addition to a mutation in VHL coding sequences) and in 1 large family with typical VHL disease but without any alteration in the other VHL exons. In this study, we show that the mutations induced a dysregulation of VHL splicing with excessive retention of E1' and were associated with a downregulation of VHL protein expression. In addition, we demonstrate a pathogenic role for synonymous mutations in VHL exon 2 that altered splicing through E2-skipping in 5 families with erythrocytosis or VHL disease. In all the studied cases, the mutations differentially affected splicing, correlating with phenotype severity. This study demonstrates that cryptic exon retention and exon skipping are new VHL alterations and reveals a novel complex splicing regulation of the VHL gene. These findings open new avenues for diagnosis and research regarding the VHL-related hypoxia-signaling pathway.


Assuntos
Éxons , Predisposição Genética para Doença , Mutação , Policitemia/genética , Processamento de RNA , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Doença de von Hippel-Lindau/genética , Adolescente , Adulto , Criança , Feminino , Heterozigoto , Humanos , Masculino , Pessoa de Meia-Idade , Linhagem , Policitemia/classificação , Policitemia/patologia , Adulto Jovem , Doença de von Hippel-Lindau/patologia
9.
Clin Genitourin Cancer ; 16(4): 266-273, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29503246

RESUMO

BACKGROUND: Clear-cell renal-cell carcinoma (ccRCC) is characterized by loss of a functional Von Hippel-Lindau (VHL) protein. We investigated the potential of 3 single nucleotide polymorphisms (SNPs) in VHL as biomarkers in metastatic ccRCC (m-ccRCC) patients treated with vascular endothelial growth factor receptor (VEGFR) tyrosine kinase inhibitors (TKIs). PATIENTS AND METHODS: We genotyped 3 VHL SNPs in 199 m-ccRCC patients: rs1642742 T > C, rs1642743 A > G, and rs1678607 C > A. Primary end points were response rate (RR), progression-free survival (PFS), and overall survival (OS) after start of first-line TKI. RR was compared with Fisher's exact test, and PFS and OS with Kaplan-Meier analysis and multivariable Cox regression. Secondary end points were association with VHL promotor hypermethylation, VHL mutation status, VHL loss of heterozygosity, ≥ 25% sarcomatoid dedifferentiation, and expression of genes implicated in angiogenesis and immunoresponse (Fisher's exact test and unpaired t tests). RESULTS: The minor alleles of rs1642742 and rs1642743, known to be in close linkage disequilibrium, were associated with poor outcome, following a recessive pattern. For the rs1642742 CC versus TT/TC genotype, OS was 11 versus 26 months (hazard ratio = 2.3; 95% confidence interval, 1.2-6.6; P = .015). For the rs1642743 GG versus AA/AG genotype, OS was 15 versus 28 months (hazard ratio = 2.6; 95% confidence interval, 1.4-5.0; P = .004). After multivariable analysis, both remained linked with poor OS (P = .018 and P = .009, respectively). There was a trend toward shorter PFS and poorer RR. Both SNPs were associated with ≥ 25% sarcomatoid dedifferentiation (P = .037 and .006, respectively). No significant results were found for rs1678607. CONCLUSION: rs1642742 and rs1642743 are candidate biomarkers for poor OS in m-ccRCC patients receiving first-line VEGFR-TKI. They are associated with higher levels of sarcomatoid dedifferentiation.


Assuntos
Carcinoma de Células Renais/tratamento farmacológico , Neoplasias Renais/tratamento farmacológico , Polimorfismo de Nucleotídeo Único , Inibidores de Proteínas Quinases/uso terapêutico , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células Renais/genética , Desdiferenciação Celular , Metilação de DNA , Feminino , Humanos , Neoplasias Renais/genética , Desequilíbrio de Ligação , Perda de Heterozigosidade , Masculino , Pessoa de Meia-Idade , Variantes Farmacogenômicos , Regiões Promotoras Genéticas , Análise de Sobrevida , Resultado do Tratamento
10.
Eur Urol ; 74(4): 474-480, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-29463434

RESUMO

BACKGROUND: Metastasectomy is routinely performed in selected patients with metastatic clear-cell renal cell carcinoma (ccRCC) as an alternative to systemic therapy. In the absence of randomized trials, the benefit and best way of patient selection remain unclear. Earlier, we described four molecular ccRCC-subtypes (ccrcc1-4) that have a prognostic and predictive value upon first-line sunitinib or pazopanib. OBJECTIVE: Assess the prognostic value of ccrcc1-4 subtypes after complete metastasectomy. (1) Compare outcomes of good-prognosis ccrccc2&3-tumors with intermediate/poor-prognosis ccrcc1&4-tumors. (2) Compare outcomes of the four subtypes separately. DESIGN, SETTING, AND PARTICIPANTS: Single-center retrospective study (1995-2017), assessing 43 ccRCC patients undergoing complete metastasectomy without systemic treatment. INTERVENTION: Molecular subtype determined with established 35-gene expression classifier. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Median disease-free survival (DFS), time to systemic therapy, cancer-specific (CSS) and overall survival (OS) from metastasectomy, estimated with Kaplan-Meier method and tested against other predictors with multivariable Cox regression. RESULTS AND LIMITATIONS: Median DFS was 23 mo for ccrcc2&3-tumors versus 9 mo for ccrcc1&4-tumors (p=0.011, hazard ratio [HR]=2.6). Median time to systemic therapy was 92 mo versus 28 mo (p=0.003, HR=3.3). Median CSS was 133 mo versus 50 mo (p<0.001, HR=2.7). Median OS was 127 mo versus 50 mo (p=0.011, HR=2.5). The classification remained independent upon multivariable analysis. Outcomes remained significantly different when comparing four subtypes separately. The intrinsic heterogeneity of expression profiles is a limitation of this approach. CONCLUSION: Even after clinical patient selection, patients with a ccrcc1- or ccrcc4-tumor are at a higher risk of relapse after complete metastasectomy. Patients with a ccrcc2- or ccrcc3-tumor usually experience a long DFS. These results need validation in a larger cohort to establish the subtypes as prognostic marker. PATIENT SUMMARY: Metastasectomy is recommended for some patients with metastatic clear-cell kidney cancer; however, we do not know who will benefit the most. We show that molecular subtypes increase the possibility to predict which patients are at risk for early relapse after metastasectomy and who may benefit more from other treatment options.


Assuntos
Carcinoma de Células Renais , Perfilação da Expressão Gênica/métodos , Perfil Genético , Neoplasias Renais , Transdução de Sinais/genética , Neoplasias das Glândulas Suprarrenais/patologia , Neoplasias das Glândulas Suprarrenais/secundário , Neoplasias das Glândulas Suprarrenais/cirurgia , Idoso , Neoplasias Ósseas/patologia , Neoplasias Ósseas/secundário , Neoplasias Ósseas/cirurgia , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/patologia , Carcinoma de Células Renais/cirurgia , Análise por Conglomerados , Intervalo Livre de Doença , Feminino , Humanos , Neoplasias Renais/genética , Neoplasias Renais/patologia , Neoplasias Renais/cirurgia , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/secundário , Neoplasias Pulmonares/cirurgia , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Avaliação de Resultados em Cuidados de Saúde , Prognóstico , Procedimentos Cirúrgicos Operatórios/efeitos adversos , Procedimentos Cirúrgicos Operatórios/classificação , Procedimentos Cirúrgicos Operatórios/métodos
11.
Clin Genitourin Cancer ; 16(3): e605-e612, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29239846

RESUMO

BACKGROUND: We previously described 4 molecular subtypes of metastatic clear cell renal cell carcinoma (mccRCC), named ccrcc1-4 (Beuselinck et al, 2015). These have both prognostic and predictive value for patients treated with first-line sunitinib, with distinctive objective response rate (ORR), progression-free survival (PFS), and overall survival (OS). The ccrcc2 and ccrcc3 tumors have the best outcomes, followed by ccrcc1 and then ccrcc4. We hypothesized that these molecular subtypes would show similar outcomes with first-line pazopanib treatment. PATIENTS AND METHODS: We classified 28 mccRCC tumors treated with pazopanib as first-line therapy, as described previously. The primary endpoints were PFS and OS from the start of pazopanib. A secondary endpoint was ORR. Because there were only 2 ccrcc3 tumors, they were pooled with the ccrcc2 tumors for outcome analysis. RESULTS: PFS was 9 months for the ccrcc2 and ccrcc3 tumors, 5 months for ccrcc1 tumors, and 3 months for the ccrcc4 tumors (P = .011). The corresponding OS duration was 69, 19, and 5 months (P = .003). The corresponding ORR was 50%, 33%, and 0%. The corresponding mean tumor size decreased by 34%, 6%, and 2% (P = .032). The ccrcc1-4 classification was a stronger predictor of outcome than the International Metastatic Renal Cell Carcinoma Database Consortium score on univariate analysis (P = .011 vs. P = .094 for PFS and P = .003 vs. .013 for OS). Both remained independent on bivariate analysis. CONCLUSION: The molecular subtypes of mccRCC are associated with outcome on pazopanib as first-line therapy. The prognostic and predictive value of the ccrcc1-4 molecular classification requires validation in prospective trials.


Assuntos
Carcinoma de Células Renais/classificação , Carcinoma de Células Renais/tratamento farmacológico , Neoplasias Renais/classificação , Neoplasias Renais/tratamento farmacológico , Pirimidinas/administração & dosagem , Sulfonamidas/administração & dosagem , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Carcinoma de Células Renais/genética , Feminino , Humanos , Neoplasias Renais/genética , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , Intervalo Livre de Progressão , Estudos Prospectivos , Pirimidinas/uso terapêutico , Estudos Retrospectivos , Sulfonamidas/uso terapêutico , Resultado do Tratamento
12.
Acta Oncol ; 57(4): 498-508, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29095068

RESUMO

OBJECTIVES: Clear-cell renal cell carcinomas (ccRCC) are characterized by hyper-vascularization and can respond to vascular endothelial growth factor receptor (VEGFR) inhibitors such as sunitinib. We aimed to study the predictive value of the expression of genes in the hypoxia induced factor (HIF) - vascular endothelial growth factor (VEGF) - VEGFR-pro-angiogenic pathway in metastatic ccRCC (m-ccRCC) patients treated with sunitinib and the correlation between the expression of these genes and the molecular ccrcc-classification, the expression of genes involved in the immune-suppressive microenvironment and Von Hippel-Lindau (VHL) - and Polybromo-1 (PBRM1) - mutational status. MATERIAL AND METHODS: m-ccRCC patients treated with sunitinib as first-line targeted therapy were included. Gene expression was studied in the primary nephrectomy sample by qRT-PCR, VHL- and PBRM1-mutational status by sequencing. Response rate by RECIST, progression-free survival (PFS) and overall survival (OS) were study endpoints. RESULTS: One hundred and four patients were included. On multivariate-analysis, HIF2A-, platelet derived growth factor receptor beta (PDGFRB)-, VEGFC-, VEGFR1- and VEGFR2-expression were correlated with PFS and HIF1A-, HIF2A-, VEGFR1- and VEGFR2-expression with OS. VEGFR2-expression showed the strongest association with outcome, being significantly correlated with all outcome parameters. HIF2A, VEGFA, VEGFR1, VEGFR2 and VEGFR3 were highly expressed in the transcriptomic ccrcc2-subtype of tumors, known to be highly sensitive to sunitinib. In the total tumor series, there was no correlation nor inverse correlation between the expression of genes involved in angiogenesis and in the immune-suppressive microenvironment. In tumors with a bi-allelic PBRM1-inactivation, HIF2A-, VEGFA-, VEGFR1- and VEGFR2-expression were higher, compared to tumors with one or two functional PBRM1-alleles. CONCLUSIONS: Intratumoral expression of genes involved in the HIF-VEGF-VEGFR-pro-angiogenic pathway, especially VEGFR2, is associated with favorable outcome on sunitinib in m-ccRCCs. Several genes involved in this pathway are upregulated in the molecular ccrcc2-subgroup, which usually responds well to sunitinib.


Assuntos
Inibidores da Angiogênese/uso terapêutico , Carcinoma de Células Renais/genética , Indóis/uso terapêutico , Neoplasias Renais/genética , Neovascularização Patológica/genética , Pirróis/uso terapêutico , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células Renais/tratamento farmacológico , Carcinoma de Células Renais/mortalidade , Intervalo Livre de Doença , Feminino , Humanos , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/mortalidade , Masculino , Pessoa de Meia-Idade , Neovascularização Patológica/tratamento farmacológico , Sunitinibe , Transcriptoma , Resultado do Tratamento
13.
Eur J Drug Metab Pharmacokinet ; 41(2): 125-38, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25465228

RESUMO

This phase I, pilot clinical study was designed to evaluate the safety and the pharmacokinetic (PK) profiles of the CIME (Metabolic Identity Card) combination of ten drugs, with a view to its use as a phenotyping cocktail. Ten healthy Caucasian subjects were orally dosed with the CIME combination (caffeine-CYP1A2, repaglinide-CYP2C8, tolbutamide-CYP2C9, omeprazole-CYP2C19, dextromethorphan-CYP2D6, midazolam-CYP3A, acetaminophen-UGT1A1, 6&9 and 2B15, digoxin-P-gp, rosuvastatin-OATP1B1&3 and memantine-active renal transport). Blood was collected over 3 days and on day 7. CIME probes and relevant metabolites were assayed by LC-MS/MS and PK parameters were calculated. Main results were: (1) good safety with reversible mild or moderate adverse effects, (2) an analytical method able to quantify simultaneously the 10 probes and the major metabolites, (3) calculation of PK parameters for all probes in general agreed with published values, and (4) identification of the low CYP2D6 metabolizer. This pilot study showed that the CIME combination was well tolerated and that its pharmacokinetics could be accurately measured in healthy volunteers. This combination can now confidently be checked for sensitivity and specificity and for lack of interaction to be validated as a phenotyping cocktail.


Assuntos
Interações Medicamentosas , Preparações Farmacêuticas/metabolismo , Administração Oral , Adulto , Sistema Enzimático do Citocromo P-450/metabolismo , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/etiologia , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/metabolismo , Feminino , Voluntários Saudáveis , Humanos , Masculino , Projetos Piloto , Adulto Jovem
14.
Oncoimmunology ; 4(12): e1049804, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26587318

RESUMO

Molecular and immune classifications powerfully predict cancer patient's survival and response to therapies. We herein describe the immune tumor microenvironment of molecular subgroups of colorectal and renal cell cancers, revealing a strong correlation between tumor subtypes and distinct immune profiles.

15.
Oral Oncol ; 51(9): 848-56, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26121938

RESUMO

OBJECTIVES: Human Papillomavirus (HPV)-related oropharyngeal squamous cell carcinoma (OSCC) patients have improved prognosis compared to other head and neck (HNSCC) cancers. Since poor prognosis is associated with tumour hypoxia, we studied whether the hypoxic response is different in HPV-related cells and tumours. MATERIAL AND METHODS: HPV-positive and -negative cells were incubated in hypoxia and analyzed by qRTPCR, western blotting and cell proliferation assays. Tumours formed by xenografting these cells in nude mice were studied by IHC. HNSCC patient samples were analyzed by unsupervised clustering of hypoxia-related gene expression, quantitative real-time PCR (qRTPCR) and immunohistochemical (IHC) detection of neo-blood vessels. RESULTS AND CONCLUSION: HPV-positive and -negative cells responded differently to hypoxia, in terms of gene expression (HIF-1α, PHD-3, GLUT-1 and VEGF-A) and cell survival. Tumour xenografts formed by HPV-positive cells had fewer hypoxic areas than those formed by HPV-negative cells. HPV related tumours were less hypoxic, expressed lower levels of hypoxia-responsive genes, and had a higher density of neo-blood vessels. HPV-related OSCC display lower tumour hypoxia, which could be linked to the distinct intrinsic abilities of HPV-positive tumour cells to adapt to hypoxia and to their better prognosis.


Assuntos
Carcinoma de Células Escamosas/metabolismo , Neoplasias Orofaríngeas/metabolismo , Infecções por Papillomavirus/metabolismo , Animais , Western Blotting , Hipóxia Celular/fisiologia , Sobrevivência Celular/fisiologia , Feminino , Expressão Gênica/fisiologia , Humanos , Masculino , Camundongos , Camundongos Nus , Reação em Cadeia da Polimerase em Tempo Real
16.
Clin Cancer Res ; 21(6): 1329-39, 2015 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-25583177

RESUMO

PURPOSE: Selecting patients with metastatic clear-cell renal cell carcinoma (m-ccRCC) who might benefit from treatment with targeted tyrosine kinase inhibitors (TKI) is a challenge. Our aim was to identify molecular markers associated with outcome in patients with m-ccRCC treated with sunitinib. EXPERIMENTAL DESIGN: We performed global transcriptome analyses on 53 primary resected ccRCC tumors from patients who developed metastatic disease and were treated with first-line sunitinib. We also determined chromosome copy-number aberrations, methylation status, and gene mutations in von Hippel-Lindau and PBRM1. Molecular data were analyzed in relation with response rate (RR), progression-free survival (PFS), and overall survival (OS). Validation was performed in 47 additional ccRCC samples treated in first-line metastatic setting with sunitinib. RESULTS: Unsupervised transcriptome analysis identified 4 robust ccRCC subtypes (ccrcc1 to 4) related to previous molecular classifications that were associated with different responses to sunitinib treatment. ccrcc1/ccrcc4 tumors had a lower RR (P = 0.005) and a shorter PFS and OS than ccrcc2/ccrcc3 tumors (P = 0.001 and 0.0003, respectively). These subtypes were the only significant covariate in the multivariate Cox model for PFS and OS (P = 0.017 and 0.006, respectively). ccrcc1/ccrcc4 tumors were characterized by a stem-cell polycomb signature and CpG hypermethylation, whereas ccrcc3 tumors, sensitive to sunitinib, did not exhibit cellular response to hypoxia. Moreover, ccrcc4 tumors exhibited sarcomatoid differentiation with a strong inflammatory, Th1-oriented but suppressive immune microenvironment, with high expression of PDCD1 (PD-1) and its ligands. CONCLUSIONS: ccRCC molecular subtypes are predictive of sunitinib response in metastatic patients, and could be used for personalized mRCC treatment with TKIs, demethylating or immunomodulatory drugs.


Assuntos
Antineoplásicos/uso terapêutico , Carcinoma de Células Renais/classificação , Carcinoma de Células Renais/tratamento farmacológico , Indóis/uso terapêutico , Neoplasias Renais/tratamento farmacológico , Pirróis/uso terapêutico , Inibidores da Angiogênese/uso terapêutico , Carcinoma de Células Renais/genética , Intervalo Livre de Doença , Resistencia a Medicamentos Antineoplásicos , Feminino , Perfilação da Expressão Gênica , Humanos , Neoplasias Renais/classificação , Neoplasias Renais/genética , Masculino , Mutação/genética , Sunitinibe , Resultado do Tratamento , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores
17.
Hepatology ; 59(6): 2228-37, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24443104

RESUMO

UNLABELLED: Fibrolamellar hepatocellular carcinoma (FLC) is a rare subtype of liver cancer occurring mostly in children and young adults. We have shown that FLC comprises two separate entities: pure (p-FLC) and mixed-FLC (m-FLC), differing in clinical presentation and course. We show that p-FLCs have a distinct gene expression signature different from that of m-FLCs, which have a signature similar to that of classical hepatocellular carcinomas. We found p-FLC profiles to be unique among 263 profiles related to diverse tumoral and nontumoral liver samples. We identified two distinct molecular subgroups of p-FLCs with different outcomes. Pathway analysis of p-FLCs revealed ERBB2 overexpression and an up-regulation of glycolysis, possibly leading to compensatory mitochondrial hyperplasia and oncocytic differentiation. Four of the sixteen genes most significantly overexpressed in p-FLCs were neuroendocrine genes: prohormone convertase 1 (PCSK1); neurotensin; delta/notch-like EGF repeat containing; and calcitonin. PCSK1 overexpression was validated by immunohistochemistry, yielding specific, diffuse staining of the protein throughout the cytoplasm, possibly corresponding to a functional form of this convertase. CONCLUSION: p-FLCs have a unique transcriptomic signature characterized by the strong expression of specific neuroendocrine genes, suggesting that these tumors may have a cellular origin different from that of HCC. Our data have implications for the use of genomic profiling for diagnosis and selection of targeted therapies in patients with p-FLC.


Assuntos
Carcinoma Hepatocelular/metabolismo , Perfilação da Expressão Gênica , Neoplasias Hepáticas/metabolismo , Pró-Proteína Convertase 1/metabolismo , Receptor ErbB-2/metabolismo , Transcrição Genética , Adulto , Calcitonina/genética , Calcitonina/metabolismo , Feminino , Genes erbB-2 , Glicólise/genética , Humanos , Masculino , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Neurotensina/genética , Neurotensina/metabolismo , Pró-Proteína Convertase 1/genética , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo
18.
Clin Cancer Res ; 19(15): 4174-84, 2013 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-23757353

RESUMO

PURPOSE: Distant metastasis after treatment is observed in about 20% of squamous cell carcinoma of the head and neck (HNSCC). In the absence of any validated robust biomarker, patients at higher risk for metastasis cannot be provided with tailored therapy. To identify prognostic HNSCC molecular subgroups and potential biomarkers, we have conducted genome-wide integrated analysis of four omic sets of data. EXPERIMENTAL DESIGN: Using state-of-the-art technologies, a core set of 45 metastasizing and 55 nonmetastasizing human papillomavirus (HPV)-unrelated HNSCC patient samples were analyzed at four different levels: gene expression (transcriptome), DNA methylation (methylome), DNA copy number (genome), and microRNA (miRNA) expression (miRNome). Molecular subgroups were identified by a model-based clustering analysis. Their clinical relevance was evaluated by survival analysis, and functional significance by pathway enrichment analysis. RESULTS: Patient subgroups selected by transcriptome, methylome, or miRNome integrated analysis are associated with shorter metastasis-free survival (MFS). A common subgroup, R1, selected by all three omic approaches, is statistically more significantly associated with MFS than any of the single omic-selected subgroups. R1 and non-R1 samples display similar DNA copy number landscapes, but more frequent chromosomal aberrations are observed in the R1 cluster (especially loss at 13q14.2-3). R1 tumors are characterized by alterations of pathways involved in cell-cell adhesion, extracellular matrix (ECM), epithelial-to-mesenchymal transition (EMT), immune response, and apoptosis. CONCLUSIONS: Integration of data across several omic profiles leads to better selection of patients at higher risk, identification of relevant molecular pathways of metastasis, and potential to discover biomarkers and drug targets.


Assuntos
Carcinoma de Células Escamosas/genética , Metilação de DNA/genética , Neoplasias de Cabeça e Pescoço/genética , MicroRNAs/genética , Metástase Neoplásica/genética , Transcriptoma/genética , Adulto , Idoso , Apoptose/genética , Biomarcadores Tumorais , Carcinoma de Células Escamosas/patologia , Variações do Número de Cópias de DNA/genética , Intervalo Livre de Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Neoplasias de Cabeça e Pescoço/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Terapia de Alvo Molecular , Metástase Neoplásica/patologia , Prognóstico , Carcinoma de Células Escamosas de Cabeça e Pescoço
19.
Int J Cancer ; 132(2): E26-36, 2013 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-22890882

RESUMO

Patients with human papillomavirus (HPV)-related oropharyngeal tumors display improved prognosis. The biological basis of this tumor phenotype is poorly understood. We investigated whether increased lymphocyte infiltrate in HPV-positive oropharyngeal squamous cell carcinomas could account for better prognosis. We previously identified, in an Affymetrix GeneChip analysis of 83 HPV-unrelated and 11 HPV-related squamous cell carcinoma of the oropharynx, several candidate genes, including CD8α and CD3ζ. Their expression was validated in this study by qRT-PCR on an independent clinical series of 144 oropharyngeal tumors. Immunohistochemical staining of tumor specimens was performed to evaluate infiltration of tumor stroma by CD8+ and CD4+ lymphocytes. The prognostic value of CD8α and CD3ζ expression levels was measured by Kaplan-Meier and Cox regression model analyses. Immune response-related signaling pathways were found to be deregulated in HPV-positive oropharyngeal tumors. Expression of CD8α, CD3ζ, granzyme K, CD28 and integrin αL RNAs was upregulated in HPV-positive lesions when compared with HPV-unrelated tumors (p < 0.05). Stroma of HPV-positive tumors was frequently and strongly infiltrated by CD8α- and CD3ζ-positive T cells. CD8α RNA expression correlated with both improved global (Kaplan-Meier; p = 0.005; Cox regression: p = 0.003) and disease-free (Cox regression: p = 0.04) survival. CD3ζ RNA expression correlated with improved overall survival (Cox regression: p = 0.024). These results suggest that an increased cytotoxic T-cell-based antitumor immune response is involved in improved prognosis of patients with HPV-positive tumors.


Assuntos
Antígenos CD8/metabolismo , Linfócitos T CD8-Positivos/imunologia , Carcinoma de Células Escamosas/imunologia , Neoplasias Orofaríngeas/imunologia , Infecções por Papillomavirus/imunologia , Alphapapillomavirus/genética , Complexo CD3/genética , Complexo CD3/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Carcinoma de Células Escamosas/mortalidade , Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas/virologia , Intervalo Livre de Doença , Feminino , Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Imunidade Celular/genética , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Análise de Sequência com Séries de Oligonucleotídeos , Proteínas Oncogênicas Virais/genética , Proteínas Oncogênicas Virais/metabolismo , Neoplasias Orofaríngeas/mortalidade , Neoplasias Orofaríngeas/patologia , Neoplasias Orofaríngeas/virologia , Infecções por Papillomavirus/mortalidade , Infecções por Papillomavirus/patologia , Infecções por Papillomavirus/virologia , Prognóstico , Modelos de Riscos Proporcionais , Análise de Regressão , Transcriptoma , Microambiente Tumoral
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