RESUMEN
BACKGROUND: NanoString technology is an innovative barcode-based system that requires less tissue than traditional techniques and can test for multiple fusion transcripts in a single reaction. The objective of this study was to determine the utility of NanoString technology in the detection of sarcoma-specific fusion transcripts in pediatric sarcomas. DESIGN: Probe pairs for the most common pediatric sarcoma fusion transcripts were designed for the assay. The NanoString assay was used to test 22 specific fusion transcripts in 45 sarcoma samples that had exhibited one of these fusion genes previously by reverse transcription polymerase chain reaction (RT-PCR). A mixture of frozen (n = 18), formalin-fixed, paraffin-embedded (FFPE) tissue (n = 23), and rapid extract template (n = 4) were used for testing. RESULTS: Each of the 22 transcripts tested was detected in at least one of the 45 tumor samples. The results of the NanoString assay were 100% concordant with the previous RT-PCR results for the tumor samples, and the technique was successful using both FFPE and rapid extract method. CONCLUSION: Multiplexed interrogation for sarcoma-specific fusion transcripts using NanoString technology is a reliable approach for molecular diagnosis of pediatric sarcomas and works well with FFPE tissues. Future work will involve validating additional sarcoma fusion transcripts as well as determining the optimal workflow for diagnostic purposes.
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Código de Barras del ADN Taxonómico , Fusión Génica/genética , Nanotecnología , Sarcoma/diagnóstico , Sondas de ADN/genética , Formaldehído , Humanos , Adhesión en Parafina , Pediatría , ARN Mensajero/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Sarcoma/clasificación , Sarcoma/genéticaRESUMEN
BACKGROUND: Mutations in the tail of histone H3 (K27M) are frequently found in pediatric midline high-grade glioma's but have rarely been reported in other malignancies. Recently, recurrent somatic nucleotide variants in histone H3 (H3 K27M) have been reported in group A posterior fossa ependymoma (EPN_PFA), an entity previously described to have no recurrent mutations. However, the true incidence of H3 K27M mutations in EPN_PFA is unknown. METHODS: In order to discern the frequency of K27M mutations in histone H3 in EPN_PFA, we analyzed 151 EPN_PFA previously profiled with genome-wide methylation arrays using a validated droplet digital PCR assay. RESULTS: We identified only 1 case out of 151 EPN_PFA harboring the K27M mutation indicating that histone mutations are extremely rare in EPN_PFA. Morphologically, this single mutated case is clearly consistent with an ependymoma, and the presence of the K27M mutation was confirmed using immunohistochemistry. DISCUSSION: K27M mutations are extremely rare in EPN_PFA. Routine evaluation of K27M mutations in EPN_PFA is of limited utility, and is unlikely to have any bearing on prognosis and/or future risk stratification.
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Neoplasias Encefálicas/genética , Ependimoma/genética , Histonas/genética , Mutación/genética , Neoplasias Encefálicas/diagnóstico por imagen , Niño , Ependimoma/diagnóstico por imagen , Histonas/metabolismo , Humanos , Lisina/genética , Imagen por Resonancia Magnética , Masculino , Metionina/genéticaRESUMEN
BACKGROUND: Thalamic gliomas are rare. The natural history is unpredictable, and the optimal management of these tumors in children is poorly defined. The aim was to identify outcomes, prognostic factors, and response to various modalities of treatment in a relatively large population of pediatric thalamic tumors from many centers within a fairly homogeneous health care system. METHODS: We performed a Canadian multicenter retrospective review of pediatric thalamic tumors presenting during the MRI era (1989-2012). Radiology and pathology were reviewed by central independent reviewers. Paraffin shavings for RNA extraction were taken and tested for fusion events involving KIAA1549:BRAF. Tumors were classified as unilateral or bithalamic based on their origin on imaging. Univariate and multivariate analyses on factors influencing survival were performed. RESULTS: Seventy-two thalamic tumors were identified from 11 institutions. Females represented 53% of the study population, and the mean age at presentation was 8.9 years. Sixty-two tumors were unilateral and 10 bithalamic. Unilateral tumors had a greater propensity to grow inferiorly towards the brainstem. These tumors were predominantly low grade in comparison to bithalamic tumors which were high-grade astrocytomas. The 5-year overall survival was 61 ± 13% for unithalamic tumors compared to 37 ± 32% for bithalamic tumors (p = 0.097). Multivariate analysis indicated tumor grade as the only significant prognostic factor for unithalamic tumors. Six unilateral tumors, all low grade, were BRAF fusion positive. CONCLUSION: Unilateral and bilateral thalamic tumors behave differently. Surgical resection is an appropriate treatment option in unilateral tumors, most of which are low grade, but outcome is not related to extent of resection (EOR). Bilateral thalamic tumors have a poorer prognosis, but the occasional patient does remarkably well. The efficacy of chemotherapy and radiotherapy has not been clearly demonstrated. Novel therapeutic approaches are required to improve the prognosis for malignant unilateral thalamic tumors and bilateral thalamic tumors.
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Astrocitoma/terapia , Neoplasias Encefálicas/terapia , Ependimoma/terapia , Tálamo , Adolescente , Astrocitoma/diagnóstico , Astrocitoma/genética , Neoplasias Encefálicas/diagnóstico , Neoplasias Encefálicas/genética , Canadá , Quimioterapia Adyuvante , Niño , Preescolar , Ependimoma/diagnóstico , Ependimoma/genética , Femenino , Glioma/genética , Glioma/terapia , Humanos , Lactante , Estimación de Kaplan-Meier , Modelos Lineales , Imagen por Resonancia Magnética , Masculino , Análisis Multivariante , Procedimientos Neuroquirúrgicos , Proteínas de Fusión Oncogénica/genética , Pronóstico , Modelos de Riesgos Proporcionales , Radioterapia Adyuvante , Estudios Retrospectivos , Resultado del TratamientoRESUMEN
PURPOSE: Molecular characterization is key to optimally diagnose and manage cancer. The complexity and cost of routine genomic analysis have unfortunately limited its use and denied many patients access to precision medicine. A possible solution is to rationalize use-creating a tiered approach to testing which uses inexpensive techniques for most patients and limits expensive testing to patients with the highest needs. Here, we tested the utility of this approach to molecularly characterize pediatric glioma in a cost- and time-sensitive manner. METHODS: We used a tiered testing pipeline of immunohistochemistry (IHC), customized fusion panels or fluorescence in situ hybridization (FISH), and targeted RNA sequencing in pediatric gliomas. Two distinct diagnostic algorithms were used for low- and high-grade gliomas (LGGs and HGGs). The percentage of driver alterations identified, associated testing costs, and turnaround time (TAT) are reported. RESULTS: The tiered approach successfully characterized 96% (95 of 99) of gliomas. For 82 LGGs, IHC, targeted fusion panel or FISH, and targeted RNA sequencing solved 35% (29 of 82), 29% (24 of 82), and 30% (25 of 82) of cases, respectively. A total of 64% (53 of 82) of samples were characterized without targeted RNA sequencing. Of 17 HGG samples, 13 were characterized by IHC and four were characterized by targeted RNA sequencing. The average cost per sample was more affordable when using the tiered approach as compared with up-front targeted RNA sequencing in LGG ($405 US dollars [USD] v $745 USD) and HGGs ($282 USD v $745 USD). The average TAT per sample was also shorter using the tiered approach (10 days for LGG, 5 days for HGG v 14 days for targeted RNA sequencing). CONCLUSION: Our tiered approach molecularly characterized 96% of samples in a cost- and time-sensitive manner. Such an approach may be feasible in neuro-oncology centers worldwide, particularly in resource-limited settings.
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Glioma , Humanos , Glioma/genética , Glioma/diagnóstico , Glioma/patología , Niño , Masculino , Preescolar , Femenino , Adolescente , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/economía , Neoplasias Encefálicas/diagnóstico , Hibridación Fluorescente in Situ/economía , Lactante , Inmunohistoquímica/economía , Recursos en Salud/economía , Análisis de Secuencia de ARN/economía , Configuración de Recursos LimitadosRESUMEN
With the success of immunotherapy in cancer, understanding the tumor immune microenvironment (TIME) has become increasingly important; however in pediatric brain tumors this remains poorly characterized. Accordingly, we developed a clinical immune-oncology gene expression assay and used it to profile a diverse range of 1382 samples with detailed clinical and molecular annotation. In low-grade gliomas we identify distinct patterns of immune activation with prognostic significance in BRAF V600E-mutant tumors. In high-grade gliomas, we observe immune activation and T-cell infiltrates in tumors that have historically been considered immune cold, as well as genomic correlates of inflammation levels. In mismatch repair deficient high-grade gliomas, we find that high tumor inflammation signature is a significant predictor of response to immune checkpoint inhibition, and demonstrate the potential for multimodal biomarkers to improve treatment stratification. Importantly, while overall patterns of immune activation are observed for histologically and genetically defined tumor types, there is significant variability within each entity, indicating that the TIME must be evaluated as an independent feature from diagnosis. In sum, in addition to the histology and molecular profile, this work underscores the importance of reporting on the TIME as an essential axis of cancer diagnosis in the era of personalized medicine.
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Neoplasias Encefálicas , Glioma , Microambiente Tumoral , Humanos , Microambiente Tumoral/inmunología , Microambiente Tumoral/genética , Neoplasias Encefálicas/inmunología , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Niño , Glioma/inmunología , Glioma/genética , Glioma/patología , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/inmunología , Femenino , Masculino , Adolescente , Regulación Neoplásica de la Expresión Génica , Pronóstico , Proteínas Proto-Oncogénicas B-raf/genética , Preescolar , Perfilación de la Expresión Génica , Inmunoterapia/métodos , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Inhibidores de Puntos de Control Inmunológico/farmacología , Mutación , Linfocitos T/inmunología , Medicina de Precisión/métodos , Linfocitos Infiltrantes de Tumor/inmunología , Relevancia ClínicaRESUMEN
High-grade diffuse glioma (HGG) is the leading cause of brain tumour death. While the genetic drivers of HGG have been well described, targeting these has thus far had little impact on survival suggesting other mechanisms are at play. Here we interrogate the alternative splicing landscape of pediatric and adult HGG through multi-omic analyses, uncovering an increased splicing burden compared with normal brain. The rate of recurrent alternative splicing in cancer drivers exceeds their mutation rate, a pattern that is recapitulated in pan-cancer analyses, and is associated with worse prognosis in HGG. We investigate potential oncogenicity by interrogating cancer pathways affected by alternative splicing in HGG; spliced cancer drivers include members of the RAS/MAPK pathway. RAS suppressor neurofibromin 1 is differentially spliced to a less active isoform in >80% of HGG downstream from REST upregulation, activating the RAS/MAPK pathway and reducing glioblastoma patient survival. Overall, our results identify non-mutagenic mechanisms by which cancers activate oncogenic pathways which need to accounted for in personalized medicine approaches.
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Neoplasias Encefálicas/genética , Glioma/genética , Oncogenes/genética , Empalme del ARN/genética , Adulto , Empalme Alternativo/genética , Animales , Secuencia de Bases , Sitios de Unión , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Niño , Cromatina/metabolismo , Exones/genética , Regulación Neoplásica de la Expresión Génica , Genes Relacionados con las Neoplasias , Glioma/patología , Humanos , Sistema de Señalización de MAP Quinasas , Ratones , Mutación/genética , Neurofibromina 1/genética , Neurofibromina 1/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteínas Represoras/metabolismo , Empalmosomas/genética , Factores de Transcripción/metabolismo , Proteínas ras/metabolismoRESUMEN
Low grade gliomas are the most frequent brain tumors in children and encompass a spectrum of histologic entities which are currently assigned World Health Organisation grades I and II. They differ substantially from their adult counterparts in both their underlying genetic alterations and in the infrequency with which they transform to higher grade tumors. Nonetheless, children with low grade glioma are a therapeutic challenge due to the heterogeneity in their clinical behavior - in particular, those with incomplete surgical resection often suffer repeat progressions with resultant morbidity and, in some cases, mortality. The identification of up-regulation of the RAS-mitogen-activated protein kinase (RAS/MAPK) pathway as a near universal feature of these tumors has led to the development of targeted therapeutics aimed at improving responses while mitigating patient morbidity. Here, we review how molecular information can help to further define the entities which fall under the umbrella of pediatric-type low-grade glioma. In doing so we discuss the specific molecular drivers of pediatric low grade glioma and how to effectively test for them, review the newest therapeutic agents and their utility in treating this disease, and propose a risk-based stratification system that considers both clinical and molecular parameters to aid clinicians in making treatment decisions.
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Neoplasias Encefálicas/genética , Glioma/genética , Astrocitoma/diagnóstico , Astrocitoma/tratamiento farmacológico , Astrocitoma/genética , Astrocitoma/patología , Neoplasias Encefálicas/diagnóstico , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/patología , Niño , Ganglioglioma/diagnóstico , Ganglioglioma/tratamiento farmacológico , Ganglioglioma/genética , Ganglioglioma/patología , Glioma/diagnóstico , Glioma/tratamiento farmacológico , Glioma/patología , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Inmunohistoquímica , Hibridación Fluorescente in Situ , Sistema de Señalización de MAP Quinasas/genética , Proteínas de la Membrana/genética , Quinasas de Proteína Quinasa Activadas por Mitógenos/antagonistas & inhibidores , Técnicas de Diagnóstico Molecular , Terapia Molecular Dirigida , Clasificación del Tumor , Neoplasias Neuroepiteliales/diagnóstico , Neoplasias Neuroepiteliales/tratamiento farmacológico , Neoplasias Neuroepiteliales/genética , Neoplasias Neuroepiteliales/patología , Patología Molecular , Reacción en Cadena de la Polimerasa , Polimorfismo de Nucleótido Simple , Inhibidores de Proteínas Quinasas/uso terapéutico , Proteínas Proto-Oncogénicas B-raf/antagonistas & inhibidores , Proteínas Proto-Oncogénicas B-raf/genética , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/antagonistas & inhibidores , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genética , Proteína 1 del Complejo de la Esclerosis Tuberosa/genética , Proteína 2 del Complejo de la Esclerosis Tuberosa/genética , Regulación hacia Arriba , Organización Mundial de la Salud , Proteínas ras/genéticaRESUMEN
Histone H3 lysine 27 (H3K27M) mutations represent the canonical oncohistone, occurring frequently in midline gliomas but also identified in haematopoietic malignancies and carcinomas. H3K27M functions, at least in part, through widespread changes in H3K27 trimethylation but its role in tumour initiation remains obscure. To address this, we created a transgenic mouse expressing H3.3K27M in diverse progenitor cell populations. H3.3K27M expression drives tumorigenesis in multiple tissues, which is further enhanced by Trp53 deletion. We find that H3.3K27M epigenetically activates a transcriptome, enriched for PRC2 and SOX10 targets, that overrides developmental and tissue specificity and is conserved between H3.3K27M-mutant mouse and human tumours. A key feature of the H3K27M transcriptome is activation of a RAS/MYC axis, which we find can be targeted therapeutically in isogenic and primary DIPG cell lines with H3.3K27M mutations, providing an explanation for the common co-occurrence of alterations in these pathways in human H3.3K27M-driven cancer. Taken together, these results show how H3.3K27M-driven transcriptome remodelling promotes tumorigenesis and will be critical for targeting cancers with these mutations.
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Neoplasias Encefálicas/genética , Epigénesis Genética , Glioma/genética , Histonas/genética , Proteínas Proto-Oncogénicas c-myc/genética , Proteínas ras/genética , Animales , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Modelos Animales de Enfermedad , Epigenómica , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Glioma/metabolismo , Glioma/patología , Histonas/metabolismo , Humanos , Lisina/genética , Lisina/metabolismo , Metilación , Ratones Noqueados , Proteínas Proto-Oncogénicas c-myc/metabolismo , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo , Proteínas ras/metabolismoRESUMEN
We performed genome-wide methylation analysis on 136 pediatric low-grade gliomas, identifying a unique cluster consisting of three tumors with oligodendroglioma-like histology, BRAF p.V600E mutations and recurrent whole chromosome gains of 7 and loss of 10. Morphologically, all showed similar features, including a diffusely infiltrative glioma composed of round nuclei with perinuclear halos, a chicken-wire pattern of branching capillaries and microcalcification. None showed astrocytic features or characteristics suggestive of high-grade tumors including necrosis or mitotic figures. All tumors harbored multiple chromosomal copy number abnormalities (>10 chromosomes altered), but none showed 1p/19q co-deletion or IDH1 p.R132H mutation. Hierarchical clustering and t-stochastic neighbor embedding analyses from DNA methylation data cluster them more closely to previously described pediatric-type low-grade gliomas and separate from adult gliomas. These tumors exhibit distinct clinical features; they are temporal lobe lesions occurring in adolescents and young adults with a prolonged history of seizures and all are alive with no recurrence (follow-up 3.2 to 13.2 years). We encountered another young adult case with quite similar pathological appearance and molecular status except for TERT promoter mutation. Although the series is small, these may represent a new category of IDH wild-type low-grade gliomas which may be confused with "molecular GBM." Further, they highlight the heterogeneity of IDH wild-type gliomas and the relatively indolent behavior of "pediatric-type" gliomas.
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Neoplasias Encefálicas/genética , Inestabilidad Cromosómica , Mutación , Oligodendroglioma/genética , Proteínas Proto-Oncogénicas B-raf/genética , Adolescente , Adulto , Neoplasias Encefálicas/patología , Niño , Femenino , Humanos , Masculino , Oligodendroglioma/patología , Adulto JovenRESUMEN
BACKGROUND: Despite increased understanding of the genetic events underlying pediatric high-grade gliomas (pHGGs), therapeutic progress is static, with poor understanding of nongenomic drivers. We therefore investigated the role of alterations in mitochondrial function and developed an effective combination therapy against pHGGs. METHODS: Mitochondrial DNA (mtDNA) copy number was measured in a cohort of 60 pHGGs. The implication of mtDNA alteration in pHGG tumorigenesis was studied and followed by an efficacy investigation using patient-derived cultures and orthotopic xenografts. RESULTS: Average mtDNA content was significantly lower in tumors versus normal brains. Decreasing mtDNA copy number in normal human astrocytes led to a markedly increased tumorigenicity in vivo. Depletion of mtDNA in pHGG cells promoted cell migration and invasion and therapeutic resistance. Shifting glucose metabolism from glycolysis to mitochondrial oxidation with the adenosine monophosphate-activated protein kinase activator AICAR (5-aminoimidazole-4-carboxamide ribonucleotide) or the pyruvate dehydrogenase kinase inhibitor dichloroacetate (DCA) significantly inhibited pHGG viability. Using DCA to shift glucose metabolism to mitochondrial oxidation and then metformin to simultaneously target mitochondrial function disrupted energy homeostasis of tumor cells, increasing DNA damage and apoptosis. The triple combination with radiation therapy, DCA and metformin led to a more potent therapeutic effect in vitro and in vivo. CONCLUSIONS: Our results suggest metabolic alterations as an onco-requisite factor of pHGG tumorigenesis. Targeting reduced mtDNA quantity represents a promising therapeutic strategy for pHGG.
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Aminoimidazol Carboxamida/análogos & derivados , Neoplasias Encefálicas/metabolismo , ADN Mitocondrial/metabolismo , Ácido Dicloroacético/farmacología , Metabolismo Energético/fisiología , Glioma/metabolismo , Ribonucleótidos/farmacología , Aminoimidazol Carboxamida/farmacología , Animales , Neoplasias Encefálicas/genética , Niño , ADN Mitocondrial/efectos de los fármacos , ADN Mitocondrial/efectos de la radiación , Metabolismo Energético/efectos de los fármacos , Metabolismo Energético/efectos de la radiación , Dosificación de Gen , Glioma/genética , Glucólisis/efectos de los fármacos , Glucólisis/efectos de la radiación , Humanos , Ratones , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: The mitogen-activated protein kinases/extracelluar signal-regulated kinases pathway is involved in cell growth and proliferation, and mutations in BRAF have made it an oncogene of interest in pediatric cancer. Previous studies found that BRAF mutations as well as KIAA1549-BRAF fusions are common in intracranial low-grade gliomas (LGGs). Fewer studies have tested for the presence of these genetic changes in spinal LGGs. The aim of this study was to better understand the prevalence of BRAF and other genetic aberrations in spinal LGG. METHODS: We retrospectively analyzed 46 spinal gliomas from patients aged 1-25 years from Children's Hospital Colorado (CHCO) and The Hospital for Sick Children (SickKids). CHCO utilized a 67-gene panel that assessed BRAF and additionally screened for other possible genetic abnormalities of interest. At SickKids, BRAF V600E was assessed by droplet digital polymerase chain reaction and immunohistochemistry. BRAF fusions were detected by fluorescence in situ hybridization, reverse transcription polymerase chain reaction, or NanoString platform. Data were correlated with clinical information. RESULTS: Of 31 samples with complete fusion analysis, 13 (42%) harbored KIAA1549-BRAF. All 13 (100%) patients with confirmed KIAA1549-BRAF survived the entirety of the study period (median [interquartile range] follow-up time: 47 months [27-85 months]) and 15 (83.3%) fusion-negative patients survived (follow-up time: 37.5 months [19.8-69.5 months]). Other mutations of interest were also identified in this patient cohort including BRAF V600E , PTPN11, H3F3A, TP53, FGFR1, and CDKN2A deletion. CONCLUSION: KIAA1549-BRAF was seen in higher frequency than BRAF V600E or other genetic aberrations in pediatric spinal LGGs and experienced lower death rates compared to KIAA1549-BRAF negative patients, although this was not statistically significant.
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BACKGROUND: Both genetic and methylation analysis have been shown to provide insight into the diagnosis and prognosis of many brain tumors. However, the implication of methylation profiling and its interaction with genetic alterations in pediatric low-grade gliomas (PLGGs) are unclear. METHODS: We performed a comprehensive analysis of PLGG with long-term clinical follow-up. In total 152 PLGGs were analyzed from a range of pathological subtypes, including 40 gangliogliomas. Complete molecular analysis was compared with genome-wide methylation data and outcome in all patients. For further analysis of specific PLGG groups, including BRAF p.V600E mutant gliomas, we compiled an additional cohort of clinically and genetically defined tumors from 3 large centers. RESULTS: Unsupervised hierarchical clustering revealed 5 novel subgroups of PLGG. These were dominated by nonneoplastic factors such as tumor location and lymphocytic infiltration. Midline PLGG clustered together while deep hemispheric lesions differed from lesions in the periphery. Mutations were distributed throughout these location-driven clusters of PLGG. A novel methylation cluster suggesting high lymphocyte infiltration was confirmed pathologically and exhibited worse progression-free survival compared with PLGG harboring similar molecular alterations (P = 0.008; multivariate analysis: P = 0.035). Although the current methylation classifier revealed low confidence in 44% of cases and failed to add information in most PLGG, it was helpful in reclassifying rare cases. The addition of histopathological and molecular information to specific methylation subgroups such as pleomorphic xanthoastrocytoma-like tumors could stratify these tumors into low and high risk (P = 0.0014). CONCLUSION: The PLGG methylome is affected by multiple nonneoplastic factors. Combined molecular and pathological analysis is key to provide additional information when methylation classification is used for PLGG in the clinical setting.
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Astrocitoma , Neoplasias Encefálicas , Glioma , Neoplasias Encefálicas/genética , Niño , Epigénesis Genética , Epigenómica , Glioma/genética , Humanos , MutaciónRESUMEN
Pediatric low-grade gliomas (pLGG) are frequently driven by genetic alterations in the RAS-mitogen-activated protein kinase (RAS/MAPK) pathway yet show unexplained variability in their clinical outcome. To address this, we characterized a cohort of >1,000 clinically annotated pLGG. Eighty-four percent of cases harbored a driver alteration, while those without an identified alteration also often exhibited upregulation of the RAS/MAPK pathway. pLGG could be broadly classified based on their alteration type. Rearrangement-driven tumors were diagnosed at a younger age, enriched for WHO grade I histology, infrequently progressed, and rarely resulted in death as compared with SNV-driven tumors. Further sub-classification of clinical-molecular correlates stratified pLGG into risk categories. These data highlight the biological and clinical differences between pLGG subtypes and opens avenues for future treatment refinement.
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Biomarcadores de Tumor/genética , Neoplasias Encefálicas/genética , Variaciones en el Número de Copia de ADN , Regulación Neoplásica de la Expresión Génica , Reordenamiento Génico , Glioma/genética , Mutación , Adolescente , Neoplasias Encefálicas/clasificación , Neoplasias Encefálicas/patología , Niño , Preescolar , Estudios de Cohortes , Femenino , Perfilación de la Expresión Génica , Glioma/clasificación , Glioma/patología , Humanos , Lactante , Recién Nacido , Masculino , Proteínas Quinasas Activadas por Mitógenos/genética , Neurofibromina 1/genética , Proteínas de Fusión Oncogénica/genética , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas ras/genéticaRESUMEN
PURPOSE: Children with pediatric gliomas harboring a BRAF V600E mutation have poor outcomes with current chemoradiotherapy strategies. Our aim was to study the role of targeted BRAF inhibition in these tumors. PATIENTS AND METHODS: We collected clinical, imaging, molecular, and outcome information from patients with BRAF V600E-mutated glioma treated with BRAF inhibition across 29 centers from multiple countries. RESULTS: Sixty-seven patients were treated with BRAF inhibition (pediatric low-grade gliomas [PLGGs], n = 56; pediatric high-grade gliomas [PHGGs], n = 11) for up to 5.6 years. Objective responses were observed in 80% of PLGGs, compared with 28% observed with conventional chemotherapy (P < .001). These responses were rapid (median, 4 months) and sustained in 86% of tumors up to 5 years while receiving therapy. After discontinuation of BRAF inhibition, 76.5% (13 of 17) of patients with PLGG experienced rapid progression (median, 2.3 months). However, upon rechallenge with BRAF inhibition, 90% achieved an objective response. Poor prognostic factors in conventional therapies, such as concomitant homozygous deletion of CDKN2A, were not associated with lack of response to BRAF inhibition. In contrast, only 36% of those with PHGG responded to BRAF inhibition, with all but one tumor progressing within 18 months. In PLGG, responses translated to 3-year progression-free survival of 49.6% (95% CI, 35.3% to 69.5%) versus 29.8% (95% CI, 20% to 44.4%) for BRAF inhibition versus chemotherapy, respectively (P = .02). CONCLUSION: Use of BRAF inhibition results in robust and durable responses in BRAF V600E-mutated PLGG. Prospective studies are required to determine long-term survival and functional outcomes with BRAF inhibitor therapy in childhood gliomas.
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The use of embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) has become commonplace in the study of neuronal development, physiology, disease modelling, and therapy development. Due to the transient nature of working with these cells, it is important to regularly confirm the cell status as a naive stem cell versus a more defined neural progenitor cell (NPC). Classically, this has been done using a panel of specific antibodies to test for the expression of transcription factors known to be observed in ESCs, but not NPCs. However, this method is both time consuming and expensive. Here, we describe the use of the NanoString nCounter system for determining the levels of expression of key transcription factors that will effectively aid in determining the state of your stem cell cultures.
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Nanotecnología/métodos , Células Madre/metabolismo , Animales , Biomarcadores/metabolismo , Sondas de ADN/metabolismo , Análisis de Datos , Células Madre Embrionarias/citología , Células Madre Embrionarias/metabolismo , Regulación de la Expresión Génica , Humanos , Células Madre Pluripotentes Inducidas/citología , Células Madre Pluripotentes Inducidas/metabolismo , Células-Madre Neurales/citología , Células-Madre Neurales/metabolismoRESUMEN
Infant gliomas have paradoxical clinical behavior compared to those in children and adults: low-grade tumors have a higher mortality rate, while high-grade tumors have a better outcome. However, we have little understanding of their biology and therefore cannot explain this behavior nor what constitutes optimal clinical management. Here we report a comprehensive genetic analysis of an international cohort of clinically annotated infant gliomas, revealing 3 clinical subgroups. Group 1 tumors arise in the cerebral hemispheres and harbor alterations in the receptor tyrosine kinases ALK, ROS1, NTRK and MET. These are typically single-events and confer an intermediate outcome. Groups 2 and 3 gliomas harbor RAS/MAPK pathway mutations and arise in the hemispheres and midline, respectively. Group 2 tumors have excellent long-term survival, while group 3 tumors progress rapidly and do not respond well to chemoradiation. We conclude that infant gliomas comprise 3 subgroups, justifying the need for specialized therapeutic strategies.
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Neoplasias Encefálicas/genética , Metilación de ADN , Epigenómica/métodos , Regulación Neoplásica de la Expresión Génica , Glioma/genética , Proteínas Tirosina Quinasas Receptoras/genética , Quinasa de Linfoma Anaplásico/genética , Quinasa de Linfoma Anaplásico/metabolismo , Neoplasias Encefálicas/clasificación , Neoplasias Encefálicas/metabolismo , Femenino , Glioma/clasificación , Glioma/metabolismo , Humanos , Lactante , Recién Nacido , Masculino , Proteínas Tirosina Quinasas/genética , Proteínas Tirosina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-met/genética , Proteínas Proto-Oncogénicas c-met/metabolismo , Proteínas Tirosina Quinasas Receptoras/metabolismo , Receptor trkA/genética , Receptor trkA/metabolismo , Análisis de Supervivencia , Secuenciación del Exoma/métodosRESUMEN
Gliomas are the most common central nervous system neoplasms affecting children and can be both high- and low-grade. Paediatric low-grade glioma may be either World Health Organization grade I or grade II. Despite being classified as grade II diffuse astrocytoma, these neoplasms arising in children are distinct clinically and molecularly from their adult counterparts. They do not tend to progress to higher grade lesions and only rarely harbour an IDH mutation. Here, we review the clinical, histologic and molecular features of paediatric grade II diffuse glioma, highlighting their diagnostic criteria, prevalence across brain locations, their most common molecular features and how to test for them, and lastly the current status of therapeutic options available for their treatment.
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Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Glioma/genética , Glioma/terapia , Terapia Molecular Dirigida , Neoplasias Encefálicas/diagnóstico , Neoplasias Encefálicas/terapia , Niño , Preescolar , Ensayos Clínicos como Asunto , Inhibidor p16 de la Quinasa Dependiente de Ciclina , Inhibidor p18 de las Quinasas Dependientes de la Ciclina/genética , Eliminación de Gen , Glioma/patología , Histonas/genética , Humanos , Isocitrato Deshidrogenasa/genética , Mutación , Estadificación de Neoplasias , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas B-raf/genética , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genética , Transactivadores/genética , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Previous studies identified recurrent fusion and duplication events in pediatric low-grade glioma (pLGG). In addition to their role in diagnosis, the presence of these events aid in dictating therapy and predicting patient survival. Clinically, BRAF alterations are most commonly identified using fluorescent in situ hybridization (FISH). However, this method is costly, labor-intensive and does not identify nonBRAF events. Here, we evaluated the NanoString nCounter gene expression system for detecting 32 of the most commonly reported fusion/duplication events in pLGG. The assay was validated on 90 pLGG samples using FISH as the gold standard and showed sensitivity and specificity of 97% and 98%, respectively. We next profiled formalin-fixed paraffin-embedded preserved biopsy specimens from 429 pLGG cases. 171 (40%) of the cases within our cohort tested positive for a fusion or duplication event contained within our panel. These events, in order of prevalence, were KIAA1549-BRAF 16;9 (89/171, 52.0%), KIAA1549-BRAF 15;9 (42/171, 24.6%), KIAA1549-BRAF 16;11 (14/171, 8.2%), FGFR1-TACC1 17;7 (13/171, 7.6%), MYBL1 duplication (5/171, 2.9%), KIAA1549-BRAF 18;10 (4/171, 2.3%), KIAA1549-BRAF 15;11 (2/171, 1.2%), FAM131B-BRAF 2;9 (1/171, 0.6%), and RNF130-BRAF 3;9 (1/171, 0.6%). This work introduces NanoString as a viable clinical replacement for the detection of fusion and duplication events in pLGG.
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Neoplasias Encefálicas/genética , Glioma/genética , Proteínas de Fusión Oncogénica/genética , Proteínas Proto-Oncogénicas B-raf/genética , Biomarcadores de Tumor/genética , Biopsia , Neoplasias Encefálicas/diagnóstico , Estudios de Cohortes , Hibridación Genómica Comparativa , Femenino , Glioma/diagnóstico , Humanos , Hibridación Fluorescente in Situ , Masculino , Mutación/genética , Pediatría , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genéticaRESUMEN
Purpose BRAF V600E is a potentially highly targetable mutation detected in a subset of pediatric low-grade gliomas (PLGGs). Its biologic and clinical effect within this diverse group of tumors remains unknown. Patients and Methods A combined clinical and genetic institutional study of patients with PLGGs with long-term follow-up was performed (N = 510). Clinical and treatment data of patients with BRAF V600E mutated PLGG (n = 99) were compared with a large international independent cohort of patients with BRAF V600E mutated-PLGG (n = 180). Results BRAF V600E mutation was detected in 69 of 405 patients (17%) with PLGG across a broad spectrum of histologies and sites, including midline locations, which are not often routinely biopsied in clinical practice. Patients with BRAF V600E PLGG exhibited poor outcomes after chemotherapy and radiation therapies that resulted in a 10-year progression-free survival of 27% (95% CI, 12.1% to 41.9%) and 60.2% (95% CI, 53.3% to 67.1%) for BRAF V600E and wild-type PLGG, respectively ( P < .001). Additional multivariable clinical and molecular stratification revealed that the extent of resection and CDKN2A deletion contributed independently to poor outcome in BRAF V600E PLGG. A similar independent role for CDKN2A and resection on outcome were observed in the independent cohort. Quantitative imaging analysis revealed progressive disease and a lack of response to conventional chemotherapy in most patients with BRAF V600E PLGG. Conclusion BRAF V600E PLGG constitutes a distinct entity with poor prognosis when treated with current adjuvant therapy.
Asunto(s)
Neoplasias Encefálicas/enzimología , Glioma/enzimología , Proteínas Proto-Oncogénicas B-raf/genética , Adolescente , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/terapia , Neoplasias del Tronco Encefálico/enzimología , Neoplasias del Tronco Encefálico/genética , Neoplasias del Tronco Encefálico/patología , Neoplasias del Tronco Encefálico/terapia , Niño , Preescolar , Estudios de Cohortes , Diencéfalo/enzimología , Diencéfalo/patología , Femenino , Glioma/genética , Glioma/patología , Glioma/terapia , Humanos , Lactante , Masculino , Mutación , Clasificación del Tumor , PronósticoRESUMEN
Paediatric brain tumours arising in the thalamus present significant diagnostic and therapeutic challenges to physicians due to their sensitive midline location. As such, genetic analysis for biomarkers to aid in the diagnosis, prognosis and treatment of these tumours is needed. Here, we identified 64 thalamic gliomas with clinical follow-up and characterized targeted genomic alterations using newly optimized droplet digital and NanoString-based assays. The median age at diagnosis was 9.25 years (range, 0.63-17.55) and median survival was 6.43 (range, 0.01-27.63) years. Our cohort contained 42 and 22 tumours reviewed as low and high grade gliomas, respectively. Five (12 %) low grade and 11 (50 %) high grade gliomas were positive for the H3F3A/HIST1H3B K27M (H3K27M) mutation. Kaplan-Meier survival analysis revealed significantly worse overall survival for patients harbouring the H3K27M mutation versus H3F3A/HIST1H3B wild type (H3WT) samples (log-rank p < 0.0001) with a median survival of 1.02 vs. 9.12 years. Mitogen-activated protein kinase (MAPK) pathway activation via BRAF or FGFR1 hotspot mutations or fusion events were detected in 44 % of patients, and was associated with long-term survival in the absence of H3K27M (log-rank p < 0.0001). Multivariate analysis demonstrated H3K27M status and high grade histology to be the most significant independent predictors of poor overall survival with hazard ratios of 6.945 and 7.721 (p < 0.0001), respectively. In contrast, MAPK pathway activation is a predictor of favourable patient outcome, although not independent of other clinical factors. Importantly, we show that low grade malignancies may harbour H3K27M mutations and that these tumours show a dismal survival compared to low grade H3WT cases. Our data strongly supports the inclusion of targeted genetic testing in childhood thalamic tumours to most accurately stratify patients into appropriate risk groups.