Pyrosequencing versus methylation-specific PCR for assessment of MGMT methylation in tumor and blood samples of glioblastoma patients

Circulating biomarkers in blood may provide an interesting alternative to risky tissue biopsies in the diagnosis and follow-up of glioblastoma patients. We have assessed MGMT methylation status in blood and tissue samples from unresected glioblastoma patients who had been included in the randomized GENOM-009 trial. Paired blood and tissue samples were assessed by methylation-specific PCR (MSP) and pyrosequencing (PYR). After establishing the minimum PYR cut-off that could yield a significant difference in overall survival, we assessed the sensitivity, specificity, positive predictive value and negative predictive value (NPV) of the analyses. Methylation could be detected in cfDNA by both MSP and PYR but with low concordance with results in tissue. Sensitivity was low for both methods (31% and 38%, respectively), while specificity was higher for MSP in blood than for PYR in plasma (96% vs 76%) and NPV was similar (56 vs 57%). Concordance of results in tissue by MSP and PYR was 84.3% (P < 0.001) and correlated with outcome. We conclude that detection of cfDNA in the blood of glioblastoma patients can be an alternative when tumor tissue is not available but methods for the detection of cfDNA in blood must improve before it can replace analysis in tumor tissue

Phase II Trial of Atezolizumab Combined With Carboplatin and Pemetrexed for Patients With Advanced Nonsquamous Non–Small-Cell Lung Cancer With Untreated Brain Metastases (Atezo-Brain, GECP17/05)

PURPOSEThe Atezo-Brain study evaluated atezolizumab combined with chemotherapy in patients with advanced non-small-cell lung cancer (NSCLC) with untreated brain metastases, a population traditionally excluded from trials.METHODSThis single-arm phase II clinical trial enrolled patients with advanced nonsquamous NSCLC with untreated brain metastases without neurologic symptoms or asymptomatic with medical treatment. Dexamethasone was allowed up to 4 mg once daily. Atezolizumab plus carboplatin and pemetrexed was given for four to six cycles followed by atezolizumab plus pemetrexed until progression for a maximum of 2 years. The primary end points were to determine the progression-free survival (PFS) rate at 12 weeks and the incidence of grade >= 3 adverse events during the first 9 weeks. Intracranial outcomes were assessed using response assessment in neuro-oncology brain metastases criteria.RESULTSForty patients were enrolled and 22 (55%) were receiving corticosteroids at baseline. The overall 12-week PFS rate was 62.2% (95% credibility interval [CrI], 47.1 to 76.2). The rate of grade 3/4 adverse events during the first 9 weeks was 27.5%. Most neurologic events were grade 1 and 2 but five patients (12.5%) experienced grade 3-4 neurologic events. With a median follow-up of 31 months, intracranial median PFS was 6.9 months and response rate was 42.7% (95% CrI, 28.1 to 57.9). Systemic median PFS was 8.9 months and response rate was 45% (95% CrI, 28.1 to 57.9). The median overall survival (OS) was 11.8 months (95% CI, 7.6 to 16.9) and the 2-year OS rate was 27.5% (95% CI, 16.6 to 45.5).CONCLUSIONAtezolizumab plus carboplatin and pemetrexed demonstrates activity in patients with advanced nonsquamous NSCLC with untreated brain metastases with an acceptable safety profile

Selinexor in Advanced, Metastatic Dedifferentiated Liposarcoma: A Multinational, Randomized, Double-Blind, Placebo-Controlled Trial

PURPOSE Antitumor activity in preclinical models and a phase I study of patients with dedifferentiated liposarcoma (DD-LPS) was observed with selinexor. We evaluated the clinical benefit of selinexor in patients with previously treated DD-LPS whose sarcoma progressed on approved agents. METHODS SEAL was a phase II-III, multicenter, randomized, double-blind, placebo-controlled study. Patients age 12 years or older with advanced DD-LPS who had received two-five lines of therapy were randomly assigned (2:1) to selinexor (60 mg) or placebo twice weekly in 6-week cycles (crossover permitted). The primary end point was progression-free survival (PFS). Patients who received at least one dose of study treatment were included for safety analysis (ClinicalTrials.gov identifier: ). RESULTS Two hundred eighty-five patients were enrolled (selinexor, n = 188; placebo, n = 97). PFS was significantly longer with selinexor versus placebo: hazard ratio (HR) 0.70 (95% CI, 0.52 to 0.95; one-sided P = .011; medians 2.8 v 2.1 months), as was time to next treatment: HR 0.50 (95% CI, 0.37 to 0.66; one-sided P < .0001; medians 5.8 v 3.2 months). With crossover, no difference was observed in overall survival. The most common treatment-emergent adverse events of any grade versus grade 3 or 4 with selinexor were nausea (151 [80.7%] v 11 [5.9]), decreased appetite (113 [60.4%] v 14 [7.5%]), and fatigue (96 [51.3%] v 12 [6.4%]). Four (2.1%) and three (3.1%) patients died in the selinexor and placebo arms, respectively. Exploratory RNA sequencing analysis identified that the absence of CALB1 expression was associated with longer PFS with selinexor compared with placebo (median 6.9 v 2.2 months; HR, 0.19; P = .001). CONCLUSION Patients with advanced, refractory DD-LPS showed improved PFS and time to next treatment with selinexor compared with placebo. Supportive care and dose reductions mitigated side effects of selinexor. Prospective validation of CALB1 expression as a predictive biomarker for selinexor in DD-LPS is warranted. (C) 2022 by American Society of Clinical Oncolog

Análisis de la metilación del promotor del gen MGMT, en muestras de tejido y sangre de pacientes con glioblastoma, mediante pirosecuenciación

Antecedentes: El glioblastoma (GB) es el glioma maligno más frecuente. Mediante el tratamiento multidisciplinar con cirugía, radio y quimioterapia se consigue una supervivencia mediana de 15 meses. La metilación del promotor del gen O-6-Metilguanina-DNA-metiltransferasa (MGMT) es un conocido factor pronóstico y predictivo de respuesta a temozolomida en estos tumores, pero su análisis de manera repetida es complejo. Las rebiopsias cerebrales comportan un riesgo para el paciente, además, las muestras de tejido obtenidas pueden no representar la heterogeneidad tumoral. Los avances en el uso de biomarcadores en biopsia líquida la están posicionando como una solución a estos inconvenientes. La determinación de la metilación de MGMT se puede llevar a cabo con diversas técnicas, siendo la PCR específica de metilación (MSP) la más utilizada, tanto en ensayos clínicos como en la práctica diaria. La pirosecuenciación es una técnica sencilla y con resultados objetivos que se ha demostrado útil para el análisis de esta alteración epigenética. En este proyecto, hemos determinado el estado de metilación del promotor de MGMT, en muestras de tejido y en ctDNA de muestras sanguíneas, utilizando las técnicas de MSP y pirosecuenciación. Metodología: Se analizó la metilación de MGMT en tejido y sangre de pacientes con GB. 81 muestras de tejido y 83 de sangre, fueron valoradas mediante MSP (CpG 74-78 y 84-87), mientras que 78 de tejido, 56 de suero y 55 de plasma, lo fueron mediante pirosecuenciación (CpG 74-78). Para la pirosecuenciación, se consideraron dos puntos de corte con relevancia clínica: aquel que dividía la cohorte en dos grupos según la máxima diferencia en supervivencia global, llamado “mejor punto de corte”, y el porcentaje mínimo a partir del cual la metilación ya tenía un impacto clínico, llamado “punto de corte mínimo”. Ambos puntos de corte tenían significación estadística. Resultados: El “mejor punto de corte” en tejido fue de 11,4%, mientras que el “punto de corte mínimo” fue de 5%. El 48,6% de casos fueron positivos para la metilación en tejido, mediante MSP y el 55,7% lo fue por pirosecuenciación. Ocho casos no metilados por MSP, se detectaron metilados por pirosecuenciación, mientras que en 3 muestras pasó a la inversa. Asimismo, 4 muestras no valorables por MSP tuvieron resultado por pirosecuenciación. En plasma, el “mejor punto de corte” para la pirosecuenciación fue 3,4%. 14,9% casos fueron metilados por MSP, mientras que el 28,6% lo fue por pirosecuenciación. El “mejor punto de corte” en suero fue de 1,6%, un valor inferior a las muestras de DNA no metilado comercial y sin correlación significativa con la supervivencia global. Por este motivo, el suero se descartó para análisis adicionales. Conclusiones: La MSP y la pirosecuenciación, una vez definido el punto de corte, son dos métodos válidos para la determinación de la metilación del promotor del gen de MGMT, en tejido de pacientes con GB. Aunque la concordancia entre ellos no es perfecta, ambos ofrecen resultados que se correlacionan con la supervivencia y la respuesta al tratamiento con agentes alquilantes. Por lo tanto, ambas técnicas son útiles para detectar pacientes que podrían beneficiarse de dichos tratamiento. Pese a que, en nuestro proyecto, se ha demostrado la presencia de ctDNA en sangre y plasma de pacientes con GB, la sensibilidad para la detección de la metilación de MGMT, con los dos métodos, es baja. El suero no es una buena fuente para la determinación de metilación de MGMT en ctDNA.Background: Glioblastoma (GB) is the most frequent malignant glioma. Methyl Guanine Methyl Transferase (MGMT) promoter methylation is a known prognostic and predictive factor of response to temozolomide in these tumors. However, repeating MGMT analysis is not always feasible. Brain rebiopsies involve risk to the patient. Moreover, tissue biopsies could not always represent the tumor heterogeneity. Advances in the use of biomarkers in liquid biopsy are positioning it as a solution to these disadvantages. MGMT methylation status analysis can be carried out through several techniques, being the methylation-specific PCR (MSP) the most used. Pyrosequencing is a simple method that has proved useful for this analysis. In this project, we have determined the MGMT promoter methylation status, in tissue samples and in ctDNA from blood samples of GB patients, by MSP and pyrosequencing. Methodology: MGMT methylation was analyzed in tissue and blood from patients diagnosed with GB. We used MSP (CpG 74-78 and 84-87) in 81 tissue samples and 83 blood samples, and pyrosequencing (CpG 74-78) in 78 tissue, 56 serum and 55 plasma samples. For pyrosequencing, two cut-off points with clinical relevance were considered: one that divided the cohort into two groups according to the maximum difference in overall survival, called "best cut-off point", and the minimum percentage from which the methylation had already a clinical impact, called the "minimum cut-off point". Both were statistical significant. Results: The "best cut-off point" in tissue was 11.4%, while the "minimum cut-off point" was 5%. The 48.6% of cases were positive for methylation in tissue, by MSP, and 55.7% by pyrosequencing. Eight unmethylated cases by MSP resulted methylated by pyrosequencing, whereas in 3 samples the results were reversed. Similarly, 4 samples that were not assessed by MSP had a result by pyrosequencing. For plasma samples, the "best cut-off" for pyrosequencing was 3.4%. 14.9% were methylated by MSP, while 28.6% were methylated by pyrosequencing. The "best cut-off" in serum was 1.6%, a value below the commercial unmethylated DNA, and it had no significant correlation with overall survival. For this reason, the serum was discarded for further analysis. Conclusions: MSP and pyrosequencing, once the cut-off point is defined, are two valid methods for the MGMT methylation status assay, in tissue of GB patients. Even thought the agreement between them is not perfect, both results correlate with survival and response to treatment with alkylating agents. Therefore, both techniques are useful. Although our project has shown the presence of ctDNA in blood and plasma of GB patients, the sensitivity to determine MGMT methylation, by either method, is low. The serum does not represent a good source to assess MGMT methytation in ctDNA

Análisis de la metilación del promotor del gen MGMT, en muestras de tejido y sangre de pacientes con glioblastoma, mediante pirosecuenciación /

Departament responsable de la tesi: Departament de Medicina.Antecedentes: El glioblastoma (GB) es el glioma maligno más frecuente. Mediante el tratamiento multidisciplinar con cirugía, radio y quimioterapia se consigue una supervivencia mediana de 15 meses. La metilación del promotor del gen O-6-Metilguanina-DNA-metiltransferasa (MGMT) es un conocido factor pronóstico y predictivo de respuesta a temozolomida en estos tumores, pero su análisis de manera repetida es complejo. Las rebiopsias cerebrales comportan un riesgo para el paciente, además, las muestras de tejido obtenidas pueden no representar la heterogeneidad tumoral. Los avances en el uso de biomarcadores en biopsia líquida la están posicionando como una solución a estos inconvenientes. La determinación de la metilación de MGMT se puede llevar a cabo con diversas técnicas, siendo la PCR específica de metilación (MSP) la más utilizada, tanto en ensayos clínicos como en la práctica diaria. La pirosecuenciación es una técnica sencilla y con resultados objetivos que se ha demostrado útil para el análisis de esta alteración epigenética.En este proyecto, hemos determinado el estado de metilación del promotor de MGMT, en muestras de tejido y en ctDNA de muestras sanguíneas, utilizando las técnicas de MSP y pirosecuenciación.Metodología: Se analizó la metilación de MGMT en tejido y sangre de pacientes con GB. 81 muestras de tejido y 83 de sangre, fueron valoradas mediante MSP (CpG 74-78 y 84-87), mientras que 78 de tejido, 56 de suero y 55 de plasma, lo fueron mediante pirosecuenciación (CpG 74-78). Para la pirosecuenciación, se consideraron dos puntos de corte con relevancia clínica: aquel que dividía la cohorte en dos grupos según la máxima diferencia en supervivencia global, llamado "mejor punto de corte", y el porcentaje mínimo a partir del cual la metilación ya tenía un impacto clínico, llamado "punto de corte mínimo". Ambos puntos de corte tenían significación estadística. Resultados: El "mejor punto de corte" en tejido fue de 11,4%, mientras que el "punto de corte mínimo" fue de 5%. El 48,6% de casos fueron positivos para la metilación en tejido, mediante MSP y el 55,7% lo fue por pirosecuenciación. Ocho casos no metilados por MSP, se detectaron metilados por pirosecuenciación, mientras que en 3 muestras pasó a la inversa. Asimismo, 4 muestras no valorables por MSP tuvieron resultado por pirosecuenciación. En plasma, el "mejor punto de corte" para la pirosecuenciación fue 3,4%. 14,9% casos fueron metilados por MSP, mientras que el 28,6% lo fue por pirosecuenciación. El "mejor punto de corte" en suero fue de 1,6%, un valor inferior a las muestras de DNA no metilado comercial y sin correlación significativa con la supervivencia global. Por este motivo, el suero se descartó para análisis adicionales. Conclusiones: La MSP y la pirosecuenciación, una vez definido el punto de corte, son dos métodos válidos para la determinación de la metilación del promotor del gen de MGMT, en tejido de pacientes con GB. Aunque la concordancia entre ellos no es perfecta, ambos ofrecen resultados que se correlacionan con la supervivencia y la respuesta al tratamiento con agentes alquilantes. Por lo tanto, ambas técnicas son útiles para detectar pacientes que podrían beneficiarse de dichos tratamiento. Pese a que, en nuestro proyecto, se ha demostrado la presencia de ctDNA en sangre y plasma de pacientes con GB, la sensibilidad para la detección de la metilación de MGMT, con los dos métodos, es baja. El suero no es una buena fuente para la determinación de metilación de MGMT en ctDNA.Background: Glioblastoma (GB) is the most frequent malignant glioma. Methyl Guanine Methyl Transferase (MGMT) promoter methylation is a known prognostic and predictive factor of response to temozolomide in these tumors. However, repeating MGMT analysis is not always feasible. Brain rebiopsies involve risk to the patient. Moreover, tissue biopsies could not always represent the tumor heterogeneity. Advances in the use of biomarkers in liquid biopsy are positioning it as a solution to these disadvantages. MGMT methylation status analysis can be carried out through several techniques, being the methylation-specific PCR (MSP) the most used. Pyrosequencing is a simple method that has proved useful for this analysis. In this project, we have determined the MGMT promoter methylation status, in tissue samples and in ctDNA from blood samples of GB patients, by MSP and pyrosequencing. Methodology: MGMT methylation was analyzed in tissue and blood from patients diagnosed with GB. We used MSP (CpG 74-78 and 84-87) in 81 tissue samples and 83 blood samples, and pyrosequencing (CpG 74-78) in 78 tissue, 56 serum and 55 plasma samples. For pyrosequencing, two cut-off points with clinical relevance were considered: one that divided the cohort into two groups according to the maximum difference in overall survival, called "best cut-off point", and the minimum percentage from which the methylation had already a clinical impact, called the "minimum cut-off point". Both were statistical significant. Results: The "best cut-off point" in tissue was 11.4%, while the "minimum cut-off point" was 5%. The 48.6% of cases were positive for methylation in tissue, by MSP, and 55.7% by pyrosequencing. Eight unmethylated cases by MSP resulted methylated by pyrosequencing, whereas in 3 samples the results were reversed. Similarly, 4 samples that were not assessed by MSP had a result by pyrosequencing. For plasma samples, the "best cut-off" for pyrosequencing was 3.4%. 14.9% were methylated by MSP, while 28.6% were methylated by pyrosequencing. The "best cut-off" in serum was 1.6%, a value below the commercial unmethylated DNA, and it had no significant correlation with overall survival. For this reason, the serum was discarded for further analysis. Conclusions: MSP and pyrosequencing, once the cut-off point is defined, are two valid methods for the MGMT methylation status assay, in tissue of GB patients. Even thought the agreement between them is not perfect, both results correlate with survival and response to treatment with alkylating agents. Therefore, both techniques are useful. Although our project has shown the presence of ctDNA in blood and plasma of GB patients, the sensitivity to determine MGMT methylation, by either method, is low. The serum does not represent a good source to assess MGMT methytation in ctDNA

Tot allò que un jove voldria saber sobre Barcelona i que mai no s'hauria imaginat que existia

Estival, Albert (realització); Cortadas, Anna (guió), Àrea de Joventut i Esports (coordinació)S'acompanya amb música de la Companyia Elèctrica Dharma.Relació de totes les activitats per a joves que, organitzades per l'Àrea de Joventut, es poden realitzar a Barcelona; música, esports, casals, dansa i vídeo. - Plans: (PM) Enric Truǫ, regidor de joventut i esports, al seu darrera porta d'entrada a l'Institut Emperador Carlos, (DP) Jove sortint d'una casa amb una bossa d'esports caminant pel carrer i arribant a la font de Canaletes. Jove posant-se roba d'esport i agafant una bicicleta. Jove entrant en bicicleta al Parc de la Ciutadella. Nois i noies corrent pel parc de la Ciutadella seguint el circuit "en forma". Jove treient-se la roba d'esport. Noi sortint del funicular de Vallvidrera, (PG) Casa Blava a Vallvidrera, (DP) Nois i noies fent diferents activitats (Int.). Noi entrant en un 4-L groc. Nois plantant una tenda de campanya, (PP) Posta de sol, (DP) Nois pintant-se les cares (int.). Nois jugant a escacs, (PP) Cares de jovesDigitalitzat pel SEDA

Targeting KRAS in Lung Cancer Beyond KRAS G12C Inhibitors : The Immune Regulatory Role of KRAS and Novel Therapeutic Strategies

Approximately 20% of lung adenocarcinomas harbor KRAS mutations, an oncogene that drives tumorigenesis and has the ability to alter the immune system and the tumor immune microenvironment. While KRAS was considered "undruggable" for decades, specific KRAS G12C covalent inhibitors have recently emerged, although their promising results are limited to a subset of patients. Several other drugs targeting KRAS activation and downstream signaling pathways are currently under investigation in early-phase clinical trials. In addition, KRAS mutations can co-exist with other mutations in significant genes in cancer (e.g., STK11 and KEAP1) which induces tumor heterogeneity and promotes different responses to therapies. This review describes the molecular characterization of KRAS mutant lung cancers from a biologic perspective to its clinical implications. We aim to summarize the tumor heterogeneity of KRAS mutant lung cancers and its immune-regulatory role, to report the efficacy achieved with current immunotherapies, and to overview the therapeutic approaches targeting KRAS mutations besides KRAS G12C inhibitor

SEOM Clinical Guideline for gastrointestinal sarcomas (GIST) (2016)

Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal neoplasms of the digestive tract, and this disease has served as a paradigmatic model for successful rational development of targeted therapies. The introduction of tyrosine kinase inhibitors with activity against KIT/PDGFRA in both localized and advanced stages has remarkably improved the survival in a disease formerly deemed resistant to all systemic therapies. The Spanish Society of Medical Oncology (SEOM) guidelines provide a multidisciplinary and updated consensus for the diagnosis and treatment of GIST patients. We strongly encourage that the managing of these patients should be performed within multidisciplinary teams in reference centers

Interference effect of food and emotional stimuli in Stroop-like tasks for children and adults with Prader-Willi Syndrome

International audienceInterference effect of food and emotional stimuli in Stroop-like tasks for children and adults with Prader-Willi Syndrome. The aim of this work was to study the way items related to food or emotion are processed by a population known to have difficulties with dietary restriction, namely individuals with Prader-Willi Syndrome (PWS). Given the presence of intellectual disability (ID) in PWS, our experiments were designed to examine whether these difficulties were specific to PWS or linked with their ID. Two modified Stroop tasks (i.e., a food version and an emotional version) were administered to seventy-four children (aged between 6 and 16 years old) divided into three groups (one with PWS, one with ID matched on age and Intellectual Quotient (IQ), and one healthy group matched on age) and to eighty-four adults (aged between 18 and 48 years old) distributed in the same three groups. For both tasks, a picture version was used for the children and a word version for the adults. For the food Stroop task, (Experiment 1), materials were composed of low or high-caloric food items and stimuli not related to food. The results show a food Stroop effect for children and adults with PWS that was absent in the group of healthy participants. Moreover, a food Stroop effect was also significant for adults with ID. For the emotional Stroop task (Experiment 2), materials were composed of negative, positive and neutral stimuli. The emotional Stroop effect was also obtained for children and adults with PWS as well as for the healthy group, but not for the age- and IQ-matched group. For the PWS groups, results show a preservation to process positive pictures for children and difficulties to process negative stimuli for both age-groups. These results suggest that people with PWS have difficulties in disengaging their attention when food stimuli are present in their environment and poorer abilities to process negative ones. These difficulties endure in adulthood