Serum DNA methylome of the colorectal cancer serrated pathway enables non-invasive detection.

The clinical relevance of the colorectal cancer serrated pathway is evident, but the screening of serrated lesions remains challenging. We aimed to characterize the serum methylome of the serrated pathway and to evaluate circulating cell-free DNA (cfDNA) methylomes as a potential source of biomarkers for the non-invasive detection of serrated lesions. We collected serum samples from individuals with serrated adenocarcinoma (SAC), traditional serrated adenomas, sessile serrated lesions, hyperplastic polyps and individuals with no colorectal findings. First, we quantified cfDNA methylation with the MethylationEPIC array. Then, we compared the methylation profiles with tissue and serum datasets. Finally, we evaluated the utility of serum cfDNA methylation biomarkers. We identified a differential methylation profile able to distinguish high-risk serrated lesions from no serrated neoplasia, showing concordance with tissue methylation from SAC and sessile serrated lesions. Serum methylation profiles are pathway-specific, clearly separating serrated lesions from conventional adenomas. The combination of ninjurin 2 (NINJ2) and glutamate-rich 1 (ERICH1) methylation discriminated high-risk serrated lesions and SAC with 91.4% sensitivity (64.4% specificity), while zinc finger protein 718 (ZNF718) methylation reported 100% sensitivity for the detection of SAC (96% specificity). This is the first study exploring the serum methylome of serrated lesions. Differential methylation of cfDNA can be used for the non-invasive detection of colorectal serrated lesions.

Hemorragia digestiva por neurosfibromas en íleon / Gastrointestinal bleeding caused by neurosfibroma of the ileum

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Aportación del bloque celular en el diagnóstico de adenopatías y masas mediastínicas o hiliares realizado por ecobroncoscopia / The Contribution of Cell Blocks in the Diagnosis of Mediastinal Masses and Hilar Adenopathy Samples From Echobronchoscopy

Introducción: La punción transbronquial guiada por ecoendoscopia permite obtener bloques celulares a partir del material de punción. Nuestro objetivo fue analizar su contribución al diagnóstico citológico convencional. Metodología: Revisión retrospectiva de las punciones por ecobroncoscopia realizadas de forma consecutiva durante 2 años con diagnóstico específico. Se analizó la capacidad de generar bloques celulares, su contribución al diagnóstico y a la realización de técnicas de inmunohistoquímica. Resultados: Se revisaron 129 muestras de lesiones correspondientes a 110 pacientes. En el 91% el diagnóstico fue de malignidad. Las lesiones puncionadas más frecuentemente fueron las adenopatías 4R (28%) y subcarinal (21%). El 72% de las muestras se procesaron como bloque celular, siendo su capacidad para realizar técnicas de inmunohistoquímica significativamente mayor a la de las muestras citológicas (52,6% vs. 14%, p < 0,0001). En 4 casos el bloque permitió un diagnóstico morfológico exclusivo (3 sarcoidosis y una metástasis de adenocarcinoma prostático) y en 3 carcinomas definir el subtipo y origen. El diagnóstico exclusivo mediante bloque celular fue significativamente más frecuente en la patología benigna que en la maligna (25% vs. 0,9%, p = 0,002). Conclusiones: La obtención de bloque celular a partir de muestras de punción por ecobroncoscopia fue del 72%. Sus principales aportaciones fueron la mejora del diagnóstico de lesiones benignas y la capacidad para realizar técnicas de inmunohistoquímica cuya contribución es esencial para la tipificación de neoplasias

Background: Cell block material from puncture can be obtained with endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) in many cases. The aim of this study was to analyze the value of additional information from cell blocks obtained with EBUS-TBNA samples from mediastinal and hilar lymph nodes and masses. Methods: Review of pathology reports with a specific diagnosis obtained from EBUS-TBNA samples of mediastinal or hilar lesions, prospectively obtained over a two-year period. The generation of cell blocks from cytology needle samples, the contribution to morphological diagnosis, and the possible use of samples for immunohistochemistry were analyzed. Results: One hundred and twenty-nine samples corresponding to 110 patients were reviewed. The diagnosis was lung cancer in 81% of cases, extrapulmonary carcinoma in 10%, sarcoidosis in 4%, lymphoma in 2.7%, and tuberculosis in 0.9%. Cell blocks could be obtained in 72% of cases. Immunohistochemistry studies on the cell blocks were significantly easier to perform than on conventional smears (52.6% vs 14%, P < 0.0001). In 4 cases, the cell block provided an exclusive morphological diagnosis (3 sarcoidosis and one metastasis from prostatic carcinoma) and in 3 carcinomas, subtype and origin could be identified. Exclusive diagnoses from the cell block were significantly more frequent in benign disease than in malignant disease (25% vs 0.9%, P = 0.002). Conclusions: Cell blocks were obtained from 72% of EBUS-TBNA diagnostic procedures. The main contributions of cell blocks to pathology examinations were the possibility of carrying out immunohistochemical staining for the better classification of neoplasms, especially extrapulmonary metastatic tumors, and the improved diagnosis of benign lesions

Viabilidad de las muestras ganglionares obtenidas por ecobroncoscopia para el estudio de alteraciones epigenéticas en pacientes con cáncer de pulmón / Viability of Lymph Node Samples Obtained by Echobronchoscopy in the Study of Epigenetic Alterations in Patients With Lung Cancer

Introducción: El diagnóstico de la afectación metastásica ganglionar en el cáncer de pulmón constituye un problema, a pesar de los avances en la estadificación. La determinación del estado de metilación en ganglios podría mejorar la capacidad de las técnicas citohistológicas para detectar afectación metastásica. Nuestro objetivo fue demostrar la viabilidad de realizar estudios de metilación en muestras ganglionares citológicas. Métodos: Estudio prospectivo que incluyó 88 pacientes con diagnóstico o alta sospecha de cáncer de pulmón no microcítico, en los que se realizó una punción citológica por ecobroncoscopia de adenopatías mediastínicas y/o hiliares. Se extrajo ADN a partir de muestras citológicas ganglionares y se realizó el tratamiento con bisulfito de sodio. Los estudios de metilación se realizaron por qPCR-MS y pirosecuenciación en los genes p16/INK4a y SHOX2. Resultados: La metodología empleada permitió obtener ADN de características óptimas/buenas en el 90% de los casos. No se observaron diferencias en la concentración de ADN respecto a la estación ganglionar ni al diagnóstico final. Los análisis por qPCR-MS y pirosecuenciación no fueron posibles en un reducido número de muestras debido a baja concentración de ADN, además de la inadecuada pureza, fragmentación y/o degradación debido al tratamiento con bisulfito de sodio. Conclusión: La cuantificación de la metilación por técnicas como qPCR-MS o pirosecuenciación en muestras ganglionares obtenidas por ecobroncoscopia resulta viable siempre y cuando se logre obtener una concentración adecuada de ADN, contribuyendo a la búsqueda de biomarcadores epigenéticos que mejoren la toma de decisiones en el cáncer de pulmón potencialmente curable en beneficio del paciente

Introduction: The diagnosis of microscopic lymph node metastasis in lung cancer is challenging despite the constant advances in tumor staging. The analysis of the methylation status of certain genes in lymph node samples could improve the diagnostic capability of conventional cyto-histological methods. The aim of this study was to demonstrate the feasibility of methylation studies using cytological lymph node samples. Methods: A prospective study including 88 patients with a diagnosis or strong suspicion of non-small cell lung cancer, in which an echobronchoscopy was performed on mediastinal or hilar lymph nodes for diagnosis and/or staging purposes. DNA was extracted from cytological lymph node samples and sodium bisulfite modification was performed. Methylation studies for p16/INK4a and SHOX2 were accomplished by MS-qPCR and pyrosequencing. Results: The methodology used in our study yielded optimal/good DNA quality in 90% of the cases. No differences in DNA concentration were observed with respect to the lymph node biopsied and final diagnosis. Methylation analyses using MS-qPCR and pyrosequencing were not possible in a small number of samples mainly due to low DNA concentration, inadequate purity, fragmentation and/or degradation as a consequence of bisulfite conversion. Conclusion: Methylation quantification using MS-qPCR and pyrosequencing of cytological lymph node samples obtained using echobronchoscopy is feasible if an appropriate DNA concentration is obtained, notably contributing to the identification of epigenetic biomarkers capable of improving decision making for the benefit of potentially curable lung cancer patients

Viability of lymph node samples obtained by echobronchoscopy in the study of epigenetic alterations in patients with lung cancer.

INTRODUCTION: The diagnosis of microscopic lymph node metastasis in lung cancer is challenging despite the constant advances in tumor staging. The analysis of the methylation status of certain genes in lymph node samples could improve the diagnostic capability of conventional cyto-histological methods. The aim of this study was to demonstrate the feasibility of methylation studies using cytological lymph node samples. METHODS: Prospective study including 88 patients with a diagnosis or strong suspicion of non-small cell lung cancer, in which an echobronchoscopy was performed on mediastinal or hilar lymph nodes for diagnostic and/or staging. DNA was extracted from cytological lymph node samples and sodium bisulfite modification was performed. Methylation studies for p16/INK4a and SHOX2 were accomplished by MS-qPCR and pyrosequencing. RESULTS: The methodology used in our study yielded optimal/good DNA quality in 90% of the cases. No differences in DNA concentration were observed with respect to the lymph node biopsied and final diagnosis. Methylation analyses using MS-qPCR and pyrosequencing were not possible in a small number of samples mainly due to low DNA concentration, inadequate purity, fragmentation and/or degradation as a consequence of bisulfite conversion. CONCLUSION: Methylation quantification using MS-qPCR and pyrosequencing of cytological lymph node samples obtained using echobronchoscopy is feasible if an appropriate DNA concentration is obtained, notably contributing to the identification of epigenetic biomarkers capable of improving decision-making for the benefit of potentially curable lung cancer patients.

The contribution of cell blocks in the diagnosis of mediastinal masses and hilar adenopathy samples from echobronchoscopy.

BACKGROUND: Cell block material from puncture can be obtained with endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) in many cases. The aim of this study was to analyse the value of additional information from cell blocks obtained with EBUS-TBNA samples from mediastinal and hilar lymph nodes and masses. METHODS: Review of pathology reports with a specific diagnosis obtained from EBUS-TBNA samples of mediastinal or hilar lesions, prospectively obtained over a two-year period. The generation of cell blocks from cytology needle samples, the contribution to morphological diagnosis, and the possible use of samples for immunohistochemistry were analysed. RESULTS: One hundred and twenty-nine samples corresponding to 110 patients were reviewed. The diagnosis was lung cancer in 81% of cases, extrapulmonary carcinoma in 10%, sarcoidosis in 4%, lymphoma in 2.7%, and tuberculosis in 0.9%. Cell blocks could be obtained in 72% of cases. Immunohistochemistry studies on the cell blocks were significantly easier to perform than on conventional smears (52.6% vs. 14%, P<.0001). In 4cases, the cell block provided an exclusive morphological diagnosis (3sarcoidosis and one metastasis from prostatic carcinoma) and in 3carcinomas, subtype and origin could be identified. Exclusive diagnoses from the cell block were significantly more frequent in benign disease than in malignant disease (25% vs 0.9%, P=.002). CONCLUSIONS: Cell blocks were obtained from 72% of EBUS-TBNA diagnostic procedures. The main contributions of cell blocks to pathology examinations were the possibility of carrying out immunohistochemical staining for the better classification of neoplasms, especially extrapulmonary metastatic tumours, and the improved diagnosis of benign lesions.