RESUMEN
BACKGROUND: Cutaneous melanoma is the deadliest skin cancer, with an increasing incidence and mortality rate. Currently, staging of patients with primary melanoma is performed using histological biomarkers such as tumor thickness and ulceration. As disruption of the epigenomic landscape is recognized as a widespread feature inherent in tumor development and progression, we aimed to identify novel biomarkers providing additional clinical information over current factors using unbiased genome-wide DNA methylation analyses. METHODS: We performed a comprehensive DNA methylation analysis during all progression stages of melanoma using Infinium HumanMethylation450 BeadChips on a discovery cohort of benign nevi (n = 14) and malignant melanoma from both primary (n = 33) and metastatic (n = 28) sites, integrating the DNA methylome with gene expression data. We validated the discovered biomarkers in three independent validation cohorts by pyrosequencing and immunohistochemistry. RESULTS: We identified and validated biomarkers for, and pathways involved in, melanoma development (e.g., HOXA9 DNA methylation) and tumor progression (e.g., TBC1D16 DNA methylation). In addition, we determined a prognostic signature with potential clinical applicability and validated PON3 DNA methylation and OVOL1 protein expression as biomarkers with prognostic information independent of tumor thickness and ulceration. CONCLUSIONS: Our data underscores the importance of epigenomic regulation in triggering metastatic dissemination through the inactivation of central cancer-related pathways. Inactivation of cell-adhesion and differentiation unleashes dissemination, and subsequent activation of inflammatory and immune system programs impairs anti-tumoral defense pathways. Moreover, we identify several markers of tumor development and progression previously unrelated to melanoma, and determined a prognostic signature with potential clinical utility.
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Metilación de ADN , ADN de Neoplasias/metabolismo , Melanoma/genética , Melanoma/fisiopatología , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/fisiopatología , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/genética , Progresión de la Enfermedad , Femenino , Humanos , Masculino , Persona de Mediana Edad , Pronóstico , Melanoma Cutáneo MalignoRESUMEN
PIK3CA, which encodes the p110α subunit of PI3K, is frequently mutated and oncogenic in breast cancer. PI3Kα inhibitors are in clinical development and despite promising early clinical activity, intrinsic resistance is frequent among patients. We have previously reported that residual downstream mTORC1 activity upon treatment with PI3Kα inhibitors drives resistance to these agents. However, the mechanism underlying this phenotype is not fully understood. Here we show that in cancer cells resistant to PI3Kα inhibition, PDK1 blockade restores sensitivity to these therapies. SGK1, which is activated by PDK1, contributes to the maintenance of residual mTORC1 activity through direct phosphorylation and inhibition of TSC2. Targeting either PDK1 or SGK1 prevents mTORC1 activation, restoring the antitumoral effects of PI3Kα inhibition in resistant cells.
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Proteínas Inmediatas-Precoces/metabolismo , Complejos Multiproteicos/metabolismo , Inhibidores de las Quinasa Fosfoinosítidos-3 , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , Animales , Fosfatidilinositol 3-Quinasa Clase I , Humanos , Diana Mecanicista del Complejo 1 de la Rapamicina , Ratones , Ratones Noqueados , Ratones Desnudos , Fosfatidilinositol 3-Quinasas/metabolismo , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora , Transducción de SeñalRESUMEN
Fatty acids (FA) modify DNA methylation in vitro, but limited information is available on whether corresponding associations exist in vivo and reflect any short-term effect of the diet. Associations between global DNA methylation and FAs were sought in blood from lactating infants (LI; n = 49) and adult males (AMM; n = 12) equally distributed across the three conventional BMI classes. AMM provided multiple samples at 2-hour intervals during 8 hours after either a single Western diet-representative meal (post-prandial samples) or no meal (fasting samples). Lipid/glucose profile, HDAC4 promoter and PDK4 5'UTR methylation were determined in AMM. Multiple regression analysis revealed that global (in LI) and both global and PDK4-specific DNA methylation (in AMM) were positively associated with eicosapentaenoic and arachidonic acid. HDAC4 methylation was inversely associated with arachidonic acid post-prandially in AMM. Global DNA methylation did not show any defined within-day pattern that would suggest a short-term response to the diet. Nonetheless, global DNA methylation was higher in normal weight subjects both post-prandially and in fasting and coincided with higher polyunsaturated relative to monounsaturated and saturated FAs. We show for the first time strong associations of DNA methylation with specific FAs in two human cohorts of distinct age, diet and postnatal development stage.
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Metilación de ADN , Ayuno/sangre , Ácidos Grasos/sangre , Histona Desacetilasas/genética , Proteínas Serina-Treonina Quinasas/genética , Proteínas Represoras/genética , Regiones no Traducidas 5' , Adulto , Ácido Araquidónico/sangre , Dieta Occidental/efectos adversos , Ácido Eicosapentaenoico/sangre , Femenino , Estudios de Asociación Genética , Humanos , Lactante , Recién Nacido , Lactancia , Masculino , Periodo Posprandial , Regiones Promotoras Genéticas , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora , Análisis de Regresión , Células THP-1RESUMEN
Abnormal fatty acid metabolism and availability are landmarks of metabolic diseases, which in turn are associated with aberrant DNA methylation profiles. To understand the role of fatty acids in disease epigenetics, we sought DNA methylation profiles specifically induced by arachidonic (AA) or oleic acid (OA) in cultured cells and compared those with published profiles of normal and diseased tissues. THP-1 monocytes were stimulated with AA or OA and analyzed using Infinium HumanMethylation450 BeadChip (Illumina) and Human Exon 1.0 ST array (Affymetrix). Data were corroborated in mouse embryonic fibroblasts. Comparisons with publicly available data were conducted by standard bioinformatics. AA and OA elicited a complex response marked by a general DNA hypermethylation and hypomethylation in the 1-200 µM range, respectively, with a maximal differential response at the 100 µM dose. The divergent response to AA and OA was prominent within the gene body of target genes, where it correlated positively with transcription. AA-induced DNA methylation profiles were similar to the corresponding profiles described for palmitic acid, atherosclerosis, diabetes, obesity, and autism, but relatively dissimilar from OA-induced profiles. Furthermore, human atherosclerosis grade-associated DNA methylation profiles were significantly enriched in AA-induced profiles. Biochemical evidence pointed to ß-oxidation, PPAR-α, and sirtuin 1 as important mediators of AA-induced DNA methylation changes. In conclusion, AA and OA exert distinct effects on the DNA methylome. The observation that AA may contribute to shape the epigenome of important metabolic diseases, supports and expands current diet-based therapeutic and preventive efforts.
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Ácido Araquidónico/metabolismo , Aterosclerosis/genética , Metilación de ADN/efectos de los fármacos , Ácido Oléico/metabolismo , Animales , Ácido Araquidónico/administración & dosificación , Aterosclerosis/metabolismo , Aterosclerosis/patología , Epigénesis Genética/efectos de los fármacos , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Genoma Humano , Humanos , Metabolismo de los Lípidos/genética , Ratones , Monocitos/efectos de los fármacos , Monocitos/metabolismo , Ácido Oléico/administración & dosificación , PPAR alfa/genéticaRESUMEN
Venous malformations (VM) are vascular malformations characterized by enlarged and distorted blood vessel channels. VM grow over time and cause substantial morbidity because of disfigurement, bleeding, and pain, representing a clinical challenge in the absence of effective treatments (Nguyenet al, 2014; Uebelhoeret al, 2012). Somatic mutations may act as drivers of these lesions, as suggested by the identification of TEK mutations in a proportion of VM (Limayeet al, 2009). We report that activating PIK3CA mutations gives rise to sporadic VM in mice, which closely resemble the histology of the human disease. Furthermore, we identified mutations in PIK3CA and related genes of the PI3K (phosphatidylinositol 3-kinase)/AKT pathway in about 30% of human VM that lack TEK alterations. PIK3CA mutations promote downstream signaling and proliferation in endothelial cells and impair normal vasculogenesis in embryonic development. We successfully treated VM in mouse models using pharmacological inhibitors of PI3Kα administered either systemically or topically. This study elucidates the etiology of a proportion of VM and proposes a therapeutic approach for this disease.
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Mutación/genética , Fosfatidilinositol 3-Quinasas/genética , Malformaciones Vasculares/enzimología , Malformaciones Vasculares/genética , Animales , Fosfatidilinositol 3-Quinasa Clase I , Embrión de Mamíferos/irrigación sanguínea , Embrión de Mamíferos/efectos de los fármacos , Embrión de Mamíferos/patología , Células Endoteliales/metabolismo , Integrasas/metabolismo , Ratones , Neovascularización Fisiológica/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Piel/irrigación sanguínea , Piel/patología , Médula Espinal/irrigación sanguínea , Médula Espinal/patología , Malformaciones Vasculares/tratamiento farmacológicoRESUMEN
The PI3K/AKT pathway is the focus of several targeted therapeutic agents for a variety of malignancies. In ERBB2-amplified breast cancer, the hyperactivation of this signaling cascade is associated with resistance to ERBB2-targeted therapy. This can occur through gain-of-function alterations or compensatory mechanisms that enter into play upon pharmacological pressure. The strong rationale in combining anti-ERBB2 agents with PI3K/AKT inhibitors, together with the identification of genomic alterations conferring sensitivity to targeted inhibition, are guiding the design of clinical studies aimed at preventing the emergence of drug resistance and achieving more durable response. In the present review, we describe the involvement of this pathway in breast cancer pathogenesis, with an emphasis on AKT kinases, and provide insight into currently available targeted agents for the treatment of ERBB2-amplified breast cancer. Finally, we provide preliminary data on a novel AKT3 mutation detected in the context of resistance to anti-ERBB2 therapy as an example of genomics-based approaches towards uncovering novel actionable targets in this setting.
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Neoplasias de la Mama/genética , Genes erbB-2/genética , Proteínas Proto-Oncogénicas c-akt/genética , Transducción de Señal/genética , Animales , Resistencia a Antineoplásicos/genética , Femenino , Humanos , Fosfatidilinositol 3-Quinasas/genéticaRESUMEN
Epigenetic changes through altered DNA methylation have been implicated in critical aspects of tumor progression, and have been extensively studied in a variety of cancer types. In contrast, our current knowledge of the aberrant genomic DNA methylation in tumor-associated fibroblasts (TAFs) or other stromal cells that act as critical coconspirators of tumor progression is very scarce. To address this gap of knowledge, we conducted genome-wide DNA methylation profiling on lung TAFs and paired control fibroblasts (CFs) from non-small cell lung cancer patients using the HumanMethylation450 microarray. We found widespread DNA hypomethylation concomitant with focal gain of DNA methylation in TAFs compared to CFs. The aberrant DNA methylation landscape of TAFs had a global impact on gene expression and a selective impact on the TGF-ß pathway. The latter included promoter hypermethylation-associated SMAD3 silencing, which was associated with hyperresponsiveness to exogenous TGF-ß1 in terms of contractility and extracellular matrix deposition. In turn, activation of CFs with exogenous TGF-ß1 partially mimicked the epigenetic alterations observed in TAFs, suggesting that TGF-ß1 may be necessary but not sufficient to elicit such alterations. Moreover, integrated pathway-enrichment analyses of the DNA methylation alterations revealed that a fraction of TAFs may be bone marrow-derived fibrocytes. Finally, survival analyses using DNA methylation and gene expression datasets identified aberrant DNA methylation on the EDARADD promoter sequence as a prognostic factor in non-small cell lung cancer patients. Our findings shed light on the unique origin and molecular alterations underlying the aberrant phenotype of lung TAFs, and identify a stromal biomarker with potential clinical relevance.
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Adenocarcinoma/genética , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Células Escamosas/genética , Metilación de ADN , Fibroblastos/metabolismo , Neoplasias Pulmonares/genética , Adenocarcinoma/patología , Carcinoma de Pulmón de Células no Pequeñas/patología , Carcinoma de Células Escamosas/patología , Epigénesis Genética , Matriz Extracelular/genética , Matriz Extracelular/metabolismo , Adhesiones Focales/genética , Adhesiones Focales/metabolismo , Regulación Neoplásica de la Expresión Génica , Humanos , Neoplasias Pulmonares/patología , Masculino , MicroARNs/genética , MicroARNs/metabolismo , Persona de Mediana Edad , Receptores de IgG/genética , Receptores de IgG/metabolismo , Análisis de Secuencia de ADN , Proteína smad3/genética , Proteína smad3/metabolismo , Transcripción Genética , Factor de Crecimiento Transformador beta1/fisiología , Células Tumorales CultivadasRESUMEN
Metastasis is responsible for most cancer-related deaths, and, among common tumor types, melanoma is one with great potential to metastasize. Here we study the contribution of epigenetic changes to the dissemination process by analyzing the changes that occur at the DNA methylation level between primary cancer cells and metastases. We found a hypomethylation event that reactivates a cryptic transcript of the Rab GTPase activating protein TBC1D16 (TBC1D16-47 kDa; referred to hereafter as TBC1D16-47KD) to be a characteristic feature of the metastatic cascade. This short isoform of TBC1D16 exacerbates melanoma growth and metastasis both in vitro and in vivo. By combining immunoprecipitation and mass spectrometry, we identified RAB5C as a new TBC1D16 target and showed that it regulates EGFR in melanoma cells. We also found that epigenetic reactivation of TBC1D16-47KD is associated with poor clinical outcome in melanoma, while conferring greater sensitivity to BRAF and MEK inhibitors.
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Progresión de la Enfermedad , Epigénesis Genética , Receptores ErbB/metabolismo , Proteínas Activadoras de GTPasa/genética , Melanoma/genética , Melanoma/patología , Animales , Línea Celular Tumoral , Metilación de ADN/efectos de los fármacos , Metilación de ADN/genética , Epigénesis Genética/efectos de los fármacos , Proteínas Activadoras de GTPasa/metabolismo , Inmunoprecipitación , Ratones Desnudos , Peso Molecular , Metástasis de la Neoplasia , Pronóstico , Regiones Promotoras Genéticas/genética , Unión Proteica/efectos de los fármacos , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Interferente Pequeño/metabolismo , Transducción de Señal/efectos de los fármacos , Activación Transcripcional/efectos de los fármacos , Activación Transcripcional/genética , Resultado del Tratamiento , Proteínas de Unión al GTP rab/metabolismoRESUMEN
OBJECTIVE: To understand whether cerebrovascular events, a major complication of atherosclerosis, are associated with any specific DNA methylation changes in the carotid plaque. APPROACH AND RESULTS: We profiled the DNA methylomes of human symptomatic carotid plaques obtained from patients who had cerebrovascular events (n=19) and asymptomatic counterparts (n=19) with a high-density microarray (≈485 000 CpG sites, Illumina), and crossed DNA methylation data with RNAseq-based expression data from an independent symptomatic carotid plaque set (n=8). Few (30) CpGs showed a significant (P<0.05; absolute Delta-Beta, >0.20) differential methylation between the 2 groups. Within symptomatic carotid plaques, DNA methylation correlated significantly with postcerebrovascular event time (range, 3-45 days; r-value range, -0.926 to 0.857; P<0.05) for ≈45 000 CpGs, the vast majority of which became hypomethylated with increasing postcerebrovascular event time. Hypomethylation was not due to erasure of the gene-body and CG-poor region hypermethylation that accompany the progression of stable lesions, but rather targeted promoters and CpG islands. Noticeably, promoter hypomethylation and increased expression of genes involved in the inhibition of the inflammatory response, defense against oxidative stress, and active DNA demethylation were observed with increasing postcerebrovascular event time. Concomitantly, histological changes consistent with phagocyte-driven plaque healing were observed. CONCLUSIONS: Weak changes in the DNA methylome distinguish symptomatic from asymptomatic plaques, but a widespread demethylation resulting in permissive transcriptional marks at atheroprotective gene promoters is established in plaques after a cerebrovascular event, thus mirroring previous observations that ruptured plaques tend to revert to a stable structure. The identified loci are candidate targets to accelerate the pace of carotid plaque stabilization.
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Amaurosis Fugax/genética , Arterias Carótidas/patología , Estenosis Carotídea/genética , Metilación de ADN , Epigénesis Genética , Ataque Isquémico Transitorio/genética , Placa Aterosclerótica , Accidente Cerebrovascular/genética , Amaurosis Fugax/diagnóstico , Enfermedades Asintomáticas , Estenosis Carotídea/complicaciones , Estenosis Carotídea/diagnóstico , Islas de CpG , Perfilación de la Expresión Génica/métodos , Regulación de la Expresión Génica , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Humanos , Ataque Isquémico Transitorio/diagnóstico , Análisis de Secuencia por Matrices de Oligonucleótidos , Fenotipo , Regiones Promotoras Genéticas , Rotura Espontánea , Accidente Cerebrovascular/diagnóstico , Transcripción GenéticaRESUMEN
BACKGROUND: Atherosclerosis severity-independent alterations in DNA methylation, a reversible and highly regulated DNA modification, have been detected in aortic atheromas, thus supporting the hypothesis that epigenetic mechanisms participate in the pathogenesis of atherosclerosis. One yet unaddressed issue is whether the progression of atherosclerosis is associated with an increase in DNA methylation drift in the vascular tissue. The purpose of the study was to identify CpG methylation profiles that vary with the progression of atherosclerosis in the human aorta. METHODS: We interrogated a set of donor-matched atherosclerotic and normal aortic samples ranging from histological grade III to VII, with a high-density (>450,000 CpG sites) DNA methylation microarray. RESULTS: We detected a correlation between histological grade and intra-pair differential methylation for 1,985 autosomal CpGs, the vast majority of which drifted towards hypermethylation with lesion progression. The identified CpG loci map to genes that are regulated by known critical transcription factors involved in atherosclerosis and participate in inflammatory and immune responses. Functional relevance was corroborated by crossing the DNA methylation profiles with expression data obtained in the same human aorta sample set, by a transcriptome-wide analysis of murine atherosclerotic aortas and from available public databases. CONCLUSIONS: Our work identifies for the first time atherosclerosis progression-specific DNA methylation profiles in the vascular tissue. These findings provide potential novel markers of lesion severity and targets to counteract the progression of the atheroma.
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Aorta/patología , Aterosclerosis/genética , Aterosclerosis/patología , Metilación de ADN , Animales , Análisis por Conglomerados , Islas de CpG , Bases de Datos Genéticas , Progresión de la Enfermedad , Epigénesis Genética , Humanos , Sistema Inmunológico , Inflamación/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Análisis de Secuencia por Matrices de Oligonucleótidos , Placa Aterosclerótica/patología , Factores de Transcripción/metabolismoRESUMEN
BACKGROUND: Epigenetic alterations may contribute to the development of atherosclerosis. In particular, DNA methylation, a reversible and highly regulated DNA modification, could influence disease onset and progression because it functions as an effector for environmental influences, including diet and lifestyle, both of which are risk factors for cardiovascular diseases. METHODS AND RESULTS: To address the role of DNA methylation changes in atherosclerosis, we compared a donor-matched healthy and atherosclerotic human aorta sample using whole-genome shotgun bisulfite sequencing. We observed that the atherosclerotic portion of the aorta was hypermethylated across many genomic loci in comparison with the matched healthy counterpart. Furthermore, we defined specific loci of differential DNA methylation using a set of donor-matched aortic samples and a high-density (>450 000 CpG sites) DNA methylation microarray. The functional importance in the disease was corroborated by crossing the DNA methylation signature with the corresponding expression data of the same samples. Among the differentially methylated CpGs associated with atherosclerosis onset, we identified genes participating in endothelial and smooth muscle functions. These findings provide new clues toward a better understanding of the molecular mechanisms of atherosclerosis. CONCLUSIONS: Our data identify an atherosclerosis-specific DNA methylation profile that highlights the contribution of different genes and pathways to the disorder. Interestingly, the observed gain of DNA methylation in the atherosclerotic lesions justifies efforts to develop DNA demethylating agents for therapeutic benefit.
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Aorta/metabolismo , Aterosclerosis/genética , Mapeo Cromosómico/métodos , Metilación de ADN , Aorta/patología , Análisis por Conglomerados , Islas de CpG , Dieta , Progresión de la Enfermedad , Epigénesis Genética , Femenino , Regulación de la Expresión Génica , Humanos , Estilo de Vida , Factores de Riesgo , Sulfitos/químicaRESUMEN
Epithelial-to-mesenchymal transition (EMT) is a plastic process in which fully differentiated epithelial cells are converted into poorly differentiated, migratory and invasive mesenchymal cells, and it has been related to the metastasis potential of tumors. This is a reversible process and cells can also eventually undergo mesenchymal-to-epithelial transition. The existence of a dynamic EMT process suggests the involvement of epigenetic shifts in the phenotype. Herein, we obtained the DNA methylomes at single-base resolution of Madin-Darby canine kidney cells undergoing EMT and translated the identified differentially methylated regions to human breast cancer cells undergoing a gain of migratory and invasive capabilities associated with the EMT phenotype. We noticed dynamic and reversible changes of DNA methylation, both on promoter sequences and gene-bodies in association with transcription regulation of EMT-related genes. Most importantly, the identified DNA methylation markers of EMT were present in primary mammary tumors in association with the epithelial or the mesenchymal phenotype of the studied breast cancer samples.
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Metilación de ADN , Transición Epitelial-Mesenquimal , Animales , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Perros , Femenino , Humanos , Células de Riñón Canino Madin DarbyRESUMEN
BACKGROUND: Pleomorphic xanthoastrocytoma (PXA) is a rare WHO grade II tumor accounting for less than 1% of all astrocytomas. Malignant transformation into PXA with anaplastic features, is unusual and correlates with poorer outcome of the patients. METHODS: Using a DNA methylation custom array, we have quantified the DNA methylation level on the promoter sequence of 807 cancer-related genes of WHO grade II (n = 11) and III PXA (n = 2) and compared to normal brain tissue (n = 10) and glioblastoma (n = 87) samples. DNA methylation levels were further confirmed on independent samples by pyrosequencing of the promoter sequences. RESULTS: Increasing DNA promoter hypermethylation events were observed in anaplastic PXA as compared with grade II samples. We further validated differential hypermethylation of CD81, HCK, HOXA5, ASCL2 and TES on anaplastic PXA and grade II tumors. Moreover, these epigenetic alterations overlap those described in glioblastoma patients, suggesting common mechanisms of tumorigenesis. CONCLUSIONS: Even taking into consideration the small size of our patient populations, our data strongly suggest that epigenome-wide profiling of PXA is a valuable tool to identify methylated genes, which may play a role in the malignant progression of PXA. These methylation alterations may provide useful biomarkers for decision-making in those patients with low-grade PXA displaying a high risk of malignant transformation.
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Astrocitoma/genética , Astrocitoma/patología , Metilación de ADN , ADN de Neoplasias/análisis , Adolescente , Adulto , Anciano , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Proteínas del Citoesqueleto/genética , Epigénesis Genética , Femenino , Regulación Neoplásica de la Expresión Génica , Proteínas de Homeodominio/genética , Humanos , Proteínas con Dominio LIM/genética , Masculino , Persona de Mediana Edad , Regiones Promotoras Genéticas , Proteínas Proto-Oncogénicas c-hck/genética , Proteínas de Unión al ARN , Análisis de Secuencia de ADN , Tetraspanina 28/genética , Adulto JovenRESUMEN
The PI3K/AKT/mTOR signaling is important for cell proliferation, survival, and metabolism. Hyperactivation of this pathway is one of the most common signaling abnormalities observed in cancer and a substantial effort has recently been made to develop molecules targeting this signaling cascade. However, it is becoming evident that PI3K inhibitors used as single agents do not elicit dramatic or durable responses. Given the numerous mechanisms mediating intrinsic and acquired resistance to these agents, hypothesis-based combinatorial strategies are probably needed to fully exploit their antitumor activity. In the first part of this review, we briefly dissect the PI3K/AKT/mTOR axis and list the most advanced compounds targeting different nodes of this cascade. The second part focuses on what we believe to be the most promising rationale-based therapeutic combinations with PI3K/AKT/mTOR inhibitors in solid tumors, with special emphasis on breast cancer.
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BACKGROUND: A major problem in cancer chemotherapy is the existence of primary resistance and/or the acquisition of secondary resistance. Many cellular defects contribute to chemoresistance, but epigenetic changes can also be a cause. METHODS: A DNA methylation microarray was used to identify epigenetic differences in oxaliplatin-sensitive and -resistant colorectal cancer cells. The candidate gene SRBC was validated by single-locus DNA methylation and expression techniques. Transfection and short hairpin experiments were used to assess oxaliplatin sensitivity. Progression-free survival (PFS) and overall survival (OS) in metastasic colorectal cancer patients were explored with Kaplan-Meier and Cox regression analyses. All statistical tests were two-sided. RESULTS: We found that oxaliplatin resistance in colorectal cancer cells depends on the DNA methylation-associated inactivation of the BRCA1 interactor SRBC gene. SRBC overexpression or depletion gives rise to sensitivity or resistance to oxaliplatin, respectively. SRBC epigenetic inactivation occurred in primary tumors from a discovery cohort of colorectal cancer patients (29.8%; n = 39 of 131), where it predicted shorter PFS (hazard ratio [HR] = 1.83; 95% confidence interval [CI] = 1.15 to 2.92; log-rank P = .01), particularly in oxaliplatin-treated case subjects for which metastasis surgery was not indicated (HR = 1.96; 95% CI = 1.13 to 3.40; log-rank P = .01). In a validation cohort of unresectable colorectal tumors treated with oxaliplatin (n = 58), SRBC hypermethylation was also associated with shorter PFS (HR = 1.90; 95% CI = 1.01 to 3.60; log-rank P = .045). CONCLUSIONS: These results provide a basis for future clinical studies to validate SRBC hypermethylation as a predictive marker for oxaliplatin resistance in colorectal cancer.
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Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Proteína BRCA1/genética , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , Resistencia a Antineoplásicos , Silenciador del Gen , Péptidos y Proteínas de Señalización Intracelular/genética , Compuestos Organoplatinos/uso terapéutico , Adulto , Anciano , Capecitabina , Línea Celular Tumoral , Quimioterapia Adyuvante , Neoplasias Colorrectales/cirugía , Islas de CpG , Metilación de ADN , Desoxicitidina/administración & dosificación , Desoxicitidina/análogos & derivados , Supervivencia sin Enfermedad , Femenino , Fluorouracilo/administración & dosificación , Fluorouracilo/análogos & derivados , Regulación Neoplásica de la Expresión Génica , Humanos , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Oxaliplatino , Modelos de Riesgos Proporcionales , ARN Interferente Pequeño/genética , TransfecciónRESUMEN
PURPOSE: Non-small-cell lung cancer (NSCLC) is a tumor in which only small improvements in clinical outcome have been achieved. The issue is critical for stage I patients for whom there are no available biomarkers that indicate which high-risk patients should receive adjuvant chemotherapy. We aimed to find DNA methylation markers that could be helpful in this regard. PATIENTS AND METHODS: A DNA methylation microarray that analyzes 450,000 CpG sites was used to study tumoral DNA obtained from 444 patients with NSCLC that included 237 stage I tumors. The prognostic DNA methylation markers were validated by a single-methylation pyrosequencing assay in an independent cohort of 143 patients with stage I NSCLC. RESULTS: Unsupervised clustering of the 10,000 most variable DNA methylation sites in the discovery cohort identified patients with high-risk stage I NSCLC who had shorter relapse-free survival (RFS; hazard ratio [HR], 2.35; 95% CI, 1.29 to 4.28; P = .004). The study in the validation cohort of the significant methylated sites from the discovery cohort found that hypermethylation of five genes was significantly associated with shorter RFS in stage I NSCLC: HIST1H4F, PCDHGB6, NPBWR1, ALX1, and HOXA9. A signature based on the number of hypermethylated events distinguished patients with high- and low-risk stage I NSCLC (HR, 3.24; 95% CI, 1.61 to 6.54; P = .001). CONCLUSION: The DNA methylation signature of NSCLC affects the outcome of stage I patients, and it can be practically determined by user-friendly polymerase chain reaction assays. The analysis of the best DNA methylation biomarkers improved prognostic accuracy beyond standard staging.
Asunto(s)
Biomarcadores de Tumor/genética , Carcinoma de Pulmón de Células no Pequeñas/genética , Metilación de ADN/genética , Neoplasias Pulmonares/genética , Transcriptoma/genética , Adulto , Anciano , Anciano de 80 o más Años , Carcinoma de Pulmón de Células no Pequeñas/mortalidad , Carcinoma de Pulmón de Células no Pequeñas/patología , Análisis por Conglomerados , Islas de CpG/genética , Supervivencia sin Enfermedad , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Estimación de Kaplan-Meier , Neoplasias Pulmonares/mortalidad , Neoplasias Pulmonares/patología , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Análisis de Secuencia por Matrices de Oligonucleótidos , Pronóstico , Modelos de Riesgos ProporcionalesRESUMEN
DNA methylation biomarkers for noninvasive diagnosis of colorectal cancer (CRC) and precursor lesions have been extensively studied. Different panels have been reported attempting to improve current protocols in clinical practice, although no definite biomarkers have been established. In the present study, we have examined patient biopsies starting from a comprehensive analysis of DNA methylation differences between paired normal and tumor samples in known cancer-related genes aiming to select the best performing candidates informative for CRC diagnosis in stool samples. Five selected markers were considered for subsequent analyses in independent biologic cohorts and in silico data sets. Among the five selected genes, three of them (AGTR1, WNT2 and SLIT2) were validated in stool DNA of affected patients with a detection sensitivity of 78% [95% confidence interval (CI), 56%-89%]. As a reference, DNA methylation of VIM and SEPT9 was evaluated in a subset of stool samples yielding sensitivities of 55% and 20%, respectively. Moreover, our panel may complement histologic and endoscopic diagnosis of inflammatory bowel disease (IBD)-associated neoplasia, as it was also efficient detecting aberrant DNA methylation in non-neoplastic tissue samples from affected patients. This novel panel of specific methylation markers can be useful for early diagnosis of CRC using stool DNA and may help in the follow-up of high-risk patients with IBD.
Asunto(s)
Biomarcadores de Tumor/genética , Neoplasias Colorrectales/diagnóstico , Metilación de ADN , ADN de Neoplasias/genética , Heces/química , Anciano , Estudios de Casos y Controles , Colon/metabolismo , Neoplasias Colorrectales/genética , Femenino , Humanos , Enfermedades Inflamatorias del Intestino/diagnóstico , Enfermedades Inflamatorias del Intestino/genética , Péptidos y Proteínas de Señalización Intercelular/genética , Masculino , Persona de Mediana Edad , Clasificación del Tumor , Estadificación de Neoplasias , Proteínas del Tejido Nervioso/genética , Pronóstico , Receptor de Angiotensina Tipo 1/genética , Recto/metabolismo , Sensibilidad y Especificidad , Proteína wnt2/genéticaRESUMEN
Using whole blood from 15 twin pairs discordant for breast cancer and high-resolution (450K) DNA methylation analysis, we identified 403 differentially methylated CpG sites including known and novel potential breast cancer genes. Confirming the results in an independent validation cohort of 21 twin pairs determined the docking protein DOK7 as a candidate for blood-based cancer diagnosis. DNA hypermethylation of the promoter region was also seen in primary breast cancer tissues and cancer cell lines. Hypermethylation of DOK7 occurs years before tumor diagnosis, suggesting a role as a powerful epigenetic blood-based biomarker as well as providing insights into breast cancer pathogenesis.
Asunto(s)
Biomarcadores/metabolismo , Neoplasias de la Mama/genética , Metilación de ADN , Epigénesis Genética , Proteínas Musculares/genética , Estudios en Gemelos como Asunto , Femenino , HumanosRESUMEN
Human aging cannot be fully understood in terms of the constrained genetic setting. Epigenetic drift is an alternative means of explaining age-associated alterations. To address this issue, we performed whole-genome bisulfite sequencing (WGBS) of newborn and centenarian genomes. The centenarian DNA had a lower DNA methylation content and a reduced correlation in the methylation status of neighboring cytosine--phosphate--guanine (CpGs) throughout the genome in comparison with the more homogeneously methylated newborn DNA. The more hypomethylated CpGs observed in the centenarian DNA compared with the neonate covered all genomic compartments, such as promoters, exonic, intronic, and intergenic regions. For regulatory regions, the most hypomethylated sequences in the centenarian DNA were present mainly at CpG-poor promoters and in tissue-specific genes, whereas a greater level of DNA methylation was observed in CpG island promoters. We extended the study to a larger cohort of newborn and nonagenarian samples using a 450,000 CpG-site DNA methylation microarray that reinforced the observation of more hypomethylated DNA sequences in the advanced age group. WGBS and 450,000 analyses of middle-age individuals demonstrated DNA methylomes in the crossroad between the newborn and the nonagenarian/centenarian groups. Our study constitutes a unique DNA methylation analysis of the extreme points of human life at a single-nucleotide resolution level.
Asunto(s)
Metilación de ADN , Anciano , Anciano de 80 o más Años , Humanos , Recién NacidoRESUMEN
Little is known about the molecular events occurring in the metastases of human tumours. Epigenetic alterations are dynamic lesions that change over the natural course of the disease, and so they might play a role in the biology of cancer cells that have departed from the primary tumour. Herein, we have adopted an epigenomic approach to identify some of these changes. Using a DNA methylation microarray platform to compare paired primary tumour and lymph node metastatic cell lines from the same patient, we observed cadherin-11 promoter CpG island hypermethylation as a likely target of the process. We found that CDH11 DNA methylation-associated transcriptional silencing occurred in the corresponding lymph node metastases of melanoma and head and neck cancer cells but not in the primary tumours. Using in vitro and in vivo cellular and mouse models for depleted or enhanced CDH11 activity, we also demonstrated that CDH11 acts as an inhibitor of tumour growth, motility and dissemination. Most importantly, the study of CDH11 5'-CpG island hypermethylation in primary tumours and lymph node metastases of cancer patients showed this epigenetic alteration to be significantly confined to the disseminated cells. Overall, these results indicate the existence of metastasis-specific epigenetic events that might contribute to the progression of the disease.