Gastrointestinal stromal tumours (GISTs) are the most common mesenchymal neoplasms of the gastrointestinal tract. The vast majority of GISTs are driven by oncogenic activation of KIT, PDGFRA or, less commonly, BRAF. Loss of succinate dehydrogenase complex activity has been identified in subsets of KIT/PDGFRA/BRAF-mutation negative tumours, yet a significant fraction of GISTs are devoid of any of such alterations. To address the pathobiology of these 'quadruple-negative' GISTs, we sought to explore the possible involvement of fusion genes. To this end we performed transcriptome sequencing on five KIT/PDGFRA/BRAF-mutation negative, SDH-proficient tumours. Intriguingly, the analysis unveiled the presence of an ETV6-NTRK3 gene fusion. The screening by FISH of 26 additional cases, including KIT/PDGFRA-mutated GISTs, failed to detect other ETV6 rearrangements beside the index case. This was a 'quadruple-negative' GIST located in the rectum, an uncommon primary site for GIST development (∼4% of all GISTs). The fusion transcript identified encompasses exon 4 of ETV6 and exon 14 of NTRK3 and therefore differs from the canonical ETV6-NTRK3 chimera of infantile fibrosarcomas. However, it retains the ability to induce IRS1 phosphorylation, activate the IGF1R downstream signalling pathway and to be targeted by IGF1R and ALK inhibitors. Thus, the ETV6-NTRK3 fusion might identify a subset of GISTs with peculiar clinicopathological characteristics which could be eligible for such therapies. Copyright © 2015 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Clinical applications of precision oncology require accurate tests that can distinguish true cancer specific mutations from errors introduced at each step of next-generation sequencing (NGS). To date, no bulk sequencing study has addressed the effects of cross-site reproducibility, nor the biological, technical and computational factors that influence variant identification. Here we report a systematic interrogation of somatic mutations in paired tumor-normal cell lines to identify factors affecting detection reproducibility and accuracy at six different centers. Using whole genome sequencing (WGS) and whole-exome sequencing (WES), we evaluated the reproducibility of different sample types with varying input amount and tumor purity, and multiple library construction protocols, followed by processing with nine bioinformatics pipelines. We found that read coverage and callers affected both WGS and WES reproducibility, but WES performance was influenced by insert fragment size, genomic copy content and the global imbalance score (GIV; G > T/C > A). Finally, taking into account library preparation protocol, tumor content, read coverage and bioinformatics processes concomitantly, we recommend actionable practices to improve the reproducibility and accuracy of NGS experiments for cancer mutation detection.

NF1 gene mutations are frequent in quadruple-negative GISTs and are often constitutional, indicating that a significant fraction of patients with apparently sporadic quadruple-negative GISTs are affected by unrecognized Neurofibromatosis Type 1 syndrome. Hence, a diagnosis of quadruple-negative GIST, especially if multifocal or with a multinodular growth pattern and a nongastric location, should alert the clinician to a possible Neurofibromatosis Type 1 syndromic condition. Clin Cancer Res; 23(1); 273-82. ©2016 AACR.

The contribution of MEF2 TFs to the tumorigenic process is still mysterious. Here we clarify that MEF2 can support both pro-oncogenic or tumor suppressive activities depending on the interaction with co-activators or co-repressors partners. Through these interactions MEF2 supervise histone modifications associated with gene activation/repression, such as H3K4 methylation and H3K27 acetylation. Critical switches for the generation of a MEF2 repressive environment are class IIa HDACs. In leiomyosarcomas (LMS), this two-faced trait of MEF2 is relevant for tumor aggressiveness. Class IIa HDACs are overexpressed in 22% of LMS, where high levels of MEF2, HDAC4 and HDAC9 inversely correlate with overall survival. The knock out of HDAC9 suppresses the transformed phenotype of LMS cells, by restoring the transcriptional proficiency of some MEF2-target loci. HDAC9 coordinates also the demethylation of H3K4me3 at the promoters of MEF2-target genes. Moreover, we show that class IIa HDACs do not bind all the regulative elements bound by MEF2. Hence, in a cell MEF2-target genes actively transcribed and strongly repressed can coexist. However, these repressed MEF2-targets are poised in terms of chromatin signature. Overall our results candidate class IIa HDACs and HDAC9 in particular, as druggable targets for a therapeutic intervention in LMS.

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Background: ␤ 2 -Microglobulin (␤2m) is a paradigmatic amyloidogenic protein. Results:In vitro, the molecular chaperone ␣B-crystallin affects the oligomerization and the fibrillogenesis of ␤2m and its R3A mutant. Conclusion: ␣B-crystallin prevents ␤2m aggregation at various stages of its aggregation pathway. Significance: Molecular chaperones may be relevant to amyloid formation in vivo.

Background: Liquid biopsy provides real-time data about prognosis and actionable mutations in metastatic breast cancer (MBC). The aim of this study was to explore the combination of circulating tumour DNA (ctDNA) analysis and circulating tumour cells (CTCs) enumeration in estimating target organs more susceptible to MBC involvement. Methods: This retrospective study analysed 88 MBC patients characterised for both CTCs and ctDNA at baseline. CTCs were isolated through the CellSearch kit, while ctDNA was analysed using the Guardant360 NGS-based assay. Sites of disease were collected on the basis of imaging. Associations were explored both through uni-and multivariate logistic regression and Fisher's exact test and the random forest machine learning algorithm. Results: After multivariate logistic regression, ESR1 mutation was the only significant factor associated with liver metastases (OR 8.10), while PIK3CA was associated with lung localisations (OR 3.74). CTC enumeration was independently associated with bone metastases (OR 10.41) and TP53 was associated with lymph node localisations (OR 2.98). The metastatic behaviour was further investigated through a random forest machine learning algorithm. Bone

Extraskeletal myxoid chondrosarcoma (EMC) is a very rare sarcoma most often arising in the soft tissue. Rare EMC of the bone have been reported. EMC exhibits distinctive clinico-pathological and genetic features; however, despite the name, it lacks any feature of cartilaginous differentiation. EMC is characterized by the rearrangement of the NR4A3, which, in most cases (about 62-75%), is fused with EWSR1 and less frequently with other partners, including TAF15 (27%), TCF12 (4%), TFG, and FUS. We herein report the identification by whole-transcriptome sequencing of HSPA8 as a novel fusion partner of NR4A3 in a case of EMC. FISH analysis confirmed the presence of a genomic HSPA8-NR4A3 translocation in the vast majority of tumor cells. Our findings expand the spectrum of NR4A3 fusion partners involved in EMC pathobiology.