Transcriptome sequencing identifies ETV6-NTRK3 as a gene fusion involved in GIST.

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.

Variant Philadelphia translocations: molecular-cytogenetic characterization and prognostic influence on frontline imatinib therapy, a GIMEMA Working Party on CML analysis.

Variant Philadelphia (Ph) chromosome translocations have been reported in 5%-10% of patients with newly diagnosed chronic myeloid leukemia (CML). Variant translocations may involve one or more chromosomes in addition to 9 and 22, and can be generated by 2 different mechanisms, 1-step and 2-step rearrangements, as revealed by fluorescence in situ hybridization. The prognostic significance of the occurrence of variant translocations has been discussed in previous studies. The European LeukemiaNet recommendations do not provide a "warning" for patients with variant translocations, but there is limited information about their outcome after therapy with tyrosine kinase inhibitors. To identify the role of variant translocations in early chronic phase (CP) CML patients treated with imatinib mesylate, we performed an analysis in a large series of 559 patients enrolled in 3 prospective imatinib trials of the Gruppo Italiano Malattie EMatologiche dell'Adulto (GIMEMA) Working Party on CML. Variant translocations occurred in 30 patients (5%). Our data show that the presence of variant translocations has no impact on the cytogenetic and molecular response or on outcome, regardless of the involvement of different mechanisms, the number of involved chromosomes, or the presence of deletions. Therefore, we suggest that patients with variant translocations do not constitute a "warning" category in the imatinib era. This study is registered at www.clinicaltrials.gov as NCT00514488 and NCT00510926.

Dual-Genotype Diffuse Low-Grade Glioma: Is It Really Time to Abandon Oligoastrocytoma As a Distinct Entity?

We report a unique case of dual-genotype oligoastrocytoma characterized by IDH2 gene mutation. The tumor was resected from the temporal lobe of a 25-year-old man. At histological examination with hematoxylin and eosin stain, it showed distinct oligodendroglial and astrocytic areas. The former retained alpha-thalassaemia/mental retardation X-linked (ATRX) immuno-expression and had absent staining for p53, while the latter had ATRX loss and p53 over-expression. Molecular analyses were separately assessed in the 2 tumor components. Gene sequencing disclosed IDH2 mutation in both, whereas oligodendroglial, but not astrocytic areas, had 1p/19q codeletion and telomerase reverse transcriptase promoter mutation. Distinction of dual-genotype oligoastrocytoma from oligodendroglioma and astrocytoma might be clinically relevant for prognosis and therapy. Because most studies that investigated the molecular phenotype of oligoastrocytomas have focused on IDH1 R132H mutated cases, we suggest further analyses on diffuse gliomas with heterogeneous (astrocytic and oligodendroglial) morphology before oligoastrocytoma is dismissed as a distinct nosological entity.

Reagent and labor cost optimization through automation of fluorescence in situ hybridization (FISH) with the VP 2000: an Italian case study.

In the modern molecular diagnostic laboratory, cost considerations are of paramount importance. Automation of complex molecular assays not only allows a laboratory to accommodate higher test volumes and throughput but also has a considerable impact on the cost of testing from the perspective of reagent costs, as well as hands-on time for skilled laboratory personnel. The following study tracked the cost of labor (hands-on time) and reagents for fluorescence in situ hybridization (FISH) testing in a routine, high-volume pathology and cytogenetics laboratory in Treviso, Italy, over a 2-y period (2011-2013). The laboratory automated FISH testing with the VP 2000 Processor, a deparaffinization, pretreatment, and special staining instrument produced by Abbott Molecular, and compared hands-on time and reagent costs to manual FISH testing. The results indicated significant cost and time saving when automating FISH with VP 2000 when more than six FISH tests were run per week. At 12 FISH assays per week, an approximate total cost reduction of 55% was observed. When running 46 FISH specimens per week, the cost saving increased to 89% versus manual testing. The results demonstrate that the VP 2000 processor can significantly reduce the cost of FISH testing in diagnostic laboratories.

Primary Synovial Sarcoma (SS) of the digestive system: a molecular and clinicopathological study of fifteen cases.

BACKGROUND: Recently a few cases of synovial sarcoma (SS) of the abdominal viscera have been reported, raising awareness about the potential for confusion between this entity and KIT-negative gastrointestinal stromal tumors (GIST). We report the clinicopathological, immunophenotypical and molecular features of fifteen more SS occurring in the stomach (8 cases), epigastric region (one case), small intestine (one case), large intestine (three cases), involving both the terminal ileum and the caecum (one case) and liver (one case). METHODS: Immunostains for SMA, DESMIN, CD34, CD117, S100, EMA, CK AE1/3, TLE1, CD56, CD99, BCL2, DOG1 were performed. Rearrangement of SS18 gene region was screened in all cases: by conventional karyotype in one case, the remaining cases were screened either by interphase FISH or Q-PCR or both. RESULTS: Ten patients were male and five female, with an age range of 17-61 years (median 44). Tumor size ranged from 2 to 15 cm (median 8). Mitoses per 10 HPF ranged from 4 to 27 (median 9.5). Eleven tumors were monophasic fibrous SS, one biphasic SS and three poorly differentiated SS. SMA, Desmin, CD34, CD117 and S100 were negative in all cases, whereas EMA and/or CK AE1/AE3 were positive in all cases. TLE1, BCL2 and CD56 were positive in all tested cases. DOG1 was positive in one case. SS18 gene region rearrangement was demonstrated in all cases. A fusion transcript was amplified in eight cases: either SS18-SSX2 or SS18-SSX1 respectively in four cases each. CONCLUSIONS: SS is increasingly recognized at visceral sites. Molecular analyses play a key role when dealing with usual histotypes in unusual sites. Correct diagnosis is crucial for appropriate therapy.

Anaplastic ependymoma of the third ventricle.

Supratentorial ependymomas are rare, especially in the third ventricle. We report the case of an ependymoma of the posterior third ventricle that was endoscopically removed just by aspiration through a flexible scope. Histologically, beside the typical pattern of growth with perivascular pseudorosettes, the tumor featured hypercellular areas with more than 10 mitoses per 10 high-power fields, consistent with grade III-anaplastic tumor. A few months later, a second neuroendoscopy offered the unique chance to appreciate the total absence of tumor tissue and the restored anatomy. However, consistently with the high grade, the tumor recurred in two different locations including the endoscopic trajectory, and spread through the cerebrospinal fluid. The patient underwent a second resective surgery and radiosurgery. Despite a cycle of chemotherapy, multiple lesions both in the ventricular system and at the level of cauda equina appeared 12 months later. A comprehensive review of intraventricular anaplastic ependymomas is also provided.