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Glioneuronal tumours are an important cause of treatment-resistant epilepsy. Subtypes of tumour are often poorly discriminated by histological features and may be difficult to diagnose due to a lack of robust diagnostic tools. This is illustrated by marked variability in the reported frequencies across different epilepsy surgical series. To address this, we used DNA methylation arrays and RNA sequencing to assay the methylation and expression profiles within a large cohort of glioneuronal tumours. By adopting a class discovery approach, we were able to identify two distinct groups of glioneuronal tumour, which only partially corresponded to the existing histological classification. Furthermore, by additional molecular analyses, we were able to identify pathogenic mutations in BRAF and FGFR1, specific to each group, in a high proportion of cases. Finally, by interrogating our expression data, we were able to show that each molecular group possessed expression phenotypes suggesting different cellular differentiation: astrocytic in one group and oligodendroglial in the second. Informed by this, we were able to identify CCND1, CSPG4, and PDGFRA as immunohistochemical targets which could distinguish between molecular groups. Our data suggest that the current histological classification of glioneuronal tumours does not adequately represent their underlying biology. Instead, we show that there are two molecular groups within glioneuronal tumours. The first of these displays astrocytic differentiation and is driven by BRAF mutations, while the second displays oligodendroglial differentiation and is driven by FGFR1 mutations.

The objective of this study is to compare two EGFR testing methodologies (a commercial real-time PCR kit and a specific EGFR mutant immunohistochemistry), with direct sequencing and to investigate the limit of detection (LOD) of both PCR-based methods. We identified EGFR mutations in 21 (16%) of the 136 tumours analyzed by direct sequencing. Interestingly, the Therascreen EGFR Mutation Test kit was able to characterize as wild-type one tumour that could not be analyzed by direct sequencing of the PCR product. We then compared the LOD of the kit and that of direct sequencing using the available mutant tumours. The kit was able to detect the presence of a mutation in a 1% dilution of the total DNA in nine of the 18 tumours (50%), which tested positive with the real-time quantitative PCR method. In all cases, EGFR mutation was identified at a dilution of 5%. Where the mutant DNA represented 30% of the total DNA, sequencing was able to detect mutations in 12 out of 19 cases (63%). Additional experiments with genetically defined standards (EGFR ΔE746-A750/+ and EGFR L858R/+) yielded similar results. Immunohistochemistry (IHC) staining with exon 19-specific antibody was seen in eight out of nine cases with E746-A750del detected by direct sequencing. Neither of the two tumours with complex deletions were positive. Of the five L858R-mutated tumours detected by the PCR methods, only two were positive for the exon 21-specific antibody. The specificity was 100% for both antibodies. The LOD of the real-time PCR method was lower than that of direct sequencing. The mutation specific IHC produced excellent specificity.

This study used Juglans regia leaves from the Gran Jefe variety; this indigenous cultivar from Nerpio is highly valued for its quality and distinct characteristics. This type of walnut is traditionally cultivated in the region and is noted for its organoleptic properties and adaptation to local climatic conditions. Two solvents were tested to determine the optimal extraction conditions for phenolic compounds: 80% ethanol and water. Direct homogenization with an Ultra-Turrax, direct ultrasound, and indirect ultrasound treatments were compared for ethanol extraction. Water extractions were conducted using direct and indirect ultrasound, infusion, and decoction. Compared to water extraction, 80% ethanol proved to be more efficient. Extracting phenolic compounds from ’Gran Jefe’ walnut leaves was most effective when using direct extraction methods without either ultrasound assistance or indirect ultrasound treatment. The main compounds identified were trans-3-caffeoylquinic acid and quercetin-3-hexoside isomer 1. The ethanolic extract obtained through direct extraction was selected to study further the bioactivities of ’Gran Jefe’ walnut leaves using C. elegans as an in vivo model. Results indicated that the leaf extract enhanced thermal and oxidative stress resistance, promoted fertility, and exhibited neuroprotective effects in models of Alzheimer’s and Parkinson’s diseases. The observed bioactivities were attributed to the free phenolics present in the ethanolic extract.

Rhabdomyosarcomas are aggressive pediatric soft‐tissue sarcomas and include high‐risk PAX3–FOXO1 fusion‐gene‐positive cases. Fibroblast growth factor receptor 4 (FGFR4) is known to contribute to rhabdomyosarcoma progression; here, we sought to investigate the involvement and potential for therapeutic targeting of other FGFRs in this disease. Cell‐based screening of FGFR inhibitors with potential for clinical repurposing (NVP‐BGJ398, nintedanib, dovitinib, and ponatinib) revealed greater sensitivity of fusion‐gene‐positive versus fusion‐gene‐negative rhabdomyosarcoma cell lines and was shown to be correlated with high expression of FGFR2 and its specific ligand, FGF7. Furthermore, patient samples exhibit higher mRNA levels of FGFR2 and FGF7 in fusion‐gene‐positive versus fusion‐gene‐negative rhabdomyosarcomas. Sustained intracellular mitogen‐activated protein kinase (MAPK) activity and FGF7 secretion into culture media during serum starvation of PAX3–FOXO1 rhabdomyosarcoma cells together with decreased cell viability after genetic silencing of FGFR2 or FGF7 was in keeping with a novel FGF7–FGFR2 autocrine loop. FGFR inhibition with NVP‐BGJ398 reduced viability and was synergistic with SN38, the active metabolite of irinotecan. In vivo, NVP‐BGJ398 abrogated xenograft growth and warrants further investigation in combination with irinotecan as a therapeutic strategy for fusion‐gene‐positive rhabdomyosarcomas. The PAX3‐FOXO1 fusion protein in high‐risk rhabdomyosarcomas increases FGFR4, FGFR2, and FGF7 levels. We demonstrate FGF7, a specific ligand of FGFR2, forms an autocrine loop that signals through MAPkinase and contributes to cell proliferation/survival. This is disrupted by the Receptor Tyrosine Kinase inhibitor (RTKi) NVP‐BGJ398, which also affects FGFR4. NVP‐BGJ398 combined with irinotecan shows potential to treat fusion‐positive rhabdomyosarcoma.

Astroblastomas are rare brain tumours which predominate in children and young adults, and have a controversial claim as a distinct entity, with no established WHO grade. Reports suggest a better outcome than high grade gliomas, though they frequently recur. Recently, they have been described to overlap with a newly-discovered group of tumours described as’high grade neuroepithelial tumour with MN1 alteration’ (CNS HGNET-MN1), defined by global methylation patterns and strongly associated with gene fusions targeting MN1 . We have studied a unique case of astroblastoma arising in a 6 year-old girl, with multiple recurrences over a period of 10 years, with the pathognomonic MN1:BEND2 fusion. Exome sequencing allowed for a phylogenetic reconstruction of tumour evolution, which when integrated with clinical, pathological and radiological data provide for a detailed understanding of disease progression, with initial treatment driving tumour dissemination along four distinct trajectories. Infiltration of distant sites was associated with a later genome doubling, whilst there was evidence of convergent evolution of different lesions acquiring distinct alterations targeting NF-κB. These data represent an unusual opportunity to understand the evolutionary history of a highly recurrent childhood brain tumour, and provide novel therapeutic targets for astroblastoma/CNS HGNET-MN1.

Fil: Pietrek, Alejandro Gerardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Bio y Geociencias del NOA. Universidad Nacional de Salta. Facultad de Ciencias Naturales. Museo de Ciencias Naturales. Instituto de Bio y Geociencias del NOA; Argentina

The construction of scientific explanations is recognised by science education researchers and curriculum developers as one of the core epistemic practices in which students should acquire proficiency. However, little is known about the knowledge and skills that teachers must and do put into practice to successfully engage their students in building explanations. Therefore, an exploratory study was conducted to analyse the instructional strategies that two secondary science teachers used to promote students’ participation in the elaboration of scientific explanations. To this end, a multi-method approach was used for data collection, which included observations and semi-structured interviews. Data analysis was carried out in two stages: the first stage was aimed at inductively encoding the participants’ interactions and discursive actions, and the second stage was aimed at performing a cross-case examination of them through constant comparative analysis. Codes were refined and clustered into seven categories. The results show that teachers quite frequently use micro strategies to interact with and guide students in explanatory episodes, but very rarely structure them into/within a complete instructional sequence to purposely promote the formulation of scientific explanations. This suggests the need to promote an explicit and conscious science teacher training to enrich their knowledge of instructional strategies for fostering and scaffolding students’ explanations.

Fil: Izquierdo, Andrea Elisa. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto Multidisciplinario de Biología Vegetal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto Multidisciplinario de Biología Vegetal; Argentina