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.

Although Gastrointestinal stromal tumors (GISTs) affect about 0.0014% of the population, GISTs smaller than 1 cm (microGISTs) are detectable in about 20% to 30% of elderly individuals. This suggests that microGISTs likely represent premalignant precursors that evolve only in a minute fraction of cases toward overt GISTs. We sought histopathologic and molecular explanations for the infrequent clinical progression in small GISTs. To investigate the mechanisms of GIST progression and identify subsets with differential malignant potential, we carried out a thorough characterization of 170 GISTs <2 cm and compared their KIT/PDGFRA status with overt GISTs. The proliferation was lower in microGISTs compared with GISTs from 1 to 2 cm (milliGISTs). In addition, microGISTs were more frequently incidental, gastric, spindle, showed an infiltrative growth pattern, a lower degree of cellularity, and abundant sclerosis. The progression was limited to 1 ileal and 1 rectal milliGISTs. KIT/PDGFRA mutations were detected in 74% of the cases. The overall frequency of KIT/PDGFRA mutation and, particularly, the frequency of KIT exon 11 mutations was significantly lower in small GISTs compared with overt GISTs. Five novel mutations, 3 in KIT (p.Phe506Leu, p.Ser692Leu, p.Glu695Lys) 2 in PDGFRA (p.Ser847X, p.Ser667Pro), plus 4 double mutations were identified. Small GISTs share with overt GIST KIT/PDGFRA mutation. Nevertheless, microGISTs display an overall lower frequency of mutations, particularly canonical KIT mutations, and also carry rare and novel mutations. These molecular features, together with the peculiar pathologic characteristics, suggest that the proliferation of these lesions is likely sustained by weakly pathogenic molecular events, supporting the epidemiologic evidence that microGISTs are self-limiting lesions.

Type II mixed cryoglobulinemia (MC) is a systemic vasculitis characterized by the presence in the serum of a monoclonal cryoprecipitable IgM with rheumatoid factor (RF) activity. Hepatitis C virus (HCV) has been recognized as its major etiologic factor. Because MC frequently evolves into overt B‐cell non‐Hodgkin's lymphoma (NHL), chronic HCV infection is hypothesized to lead to both benign and malignant lymphoproliferative disease. In this study, we investigated mutations in the V_H and V_K genes of the B‐cell clone originating the overt B‐cell lymphoma in a subject with MC. Mutational patterns were analyzed longitudinally in two bone marrow biopsies obtained at the stage of MC, as well as in multiple involved tissues (bone marrow, liver, and peripheral blood cells) at the stage of overt NHL. Hybridization of variable‐diversity‐joining (VDJ) PCR products with a probe specific for the neoplastic clone indicated that the lymphoma originated from one of the clones over‐stimulated during MC. This clone producing an IgM highly homologous to a protein with RF specificity may explain the MC syndrome in the patient. Moreover, the presence of an IgH ongoing mutation process and the expression of an Ig antigen receptor significantly homologous to an anti‐HCV protein support the hypothesis that the MC syndrome and the subsequent evolution to NHL are antigen‐driven lymphoproliferative processes possibly sustained by HCV. Furthermore, the marked reduction in intra‐clonal diversity in the last bone marrow biopsy obtained at the stage of overt NHL points out a minor dependence of the cells on the antigen‐driven mechanism, although an intrinsic propensity of the neoplastic cell to undergo replacement mutations cannot be excluded. Int. J. Cancer 87:211–216, 2000.

Objective Type II mixed cryoglobulinemia (type II MC) is often characterized by features of indolent B cell lymphoma (IBCL) found on pathologic examination of bone marrow, whereas the clinical evidence does not indicate a neoplastic disorder. To better address the issue of indolent malignant versus nonmalignant bone marrow lymphoproliferation underlying type II MC, molecular analyses of B cell clonality were performed in the present study, in conjunction with clinical and pathologic characterization. Methods Polymerase chain reaction DNA amplification of immunoglobulin heavy chain genes was performed in bone marrow biopsy specimens obtained from 15 selected patients with type II MC, all infected with hepatitis C virus. Five of them had also developed overt B cell lymphoma (OBCL) during followup. Bone marrow features were consistent with IBCL in 9 of the 15 patients (group 1) and with reactive lymphoplasmacytosis in 6 of the 15 (group 2). Results An oligoclonal B cell expansion was detected in 6 of 9 baseline bone marrow lesions from group 1 patients (biclonal or monoclonal expansion in the remaining 3 cases), and in 6 of 6 from group 2 patients. OBCL was always monoclonal. Selected lesions were analyzed by clonospecific hybridization and by cloning and sequence analysis in patients who had developed OBCL at followup. In 4 of 5 cases, OBCL did not originate from the dominant B cell clones that were overexpanded in the putative neoplastic baseline bone marrow lesions. OBCL clones showed significant homology with rheumatoid factor database sequences. Conclusion Based on the present results, as well as on evidence from previous studies of liver lesions, oligoclonal non‐neoplastic B cell proliferation in the course of chronic infection–related inflammation appears to be the key feature of type II MC. Of note, molecular evidence from target tissues supports the clinical findings both at the time of type II MC diagnosis and in cases of OBCL complication. Bone marrow pathologic findings resembling those of IBCL should thus be considered in the light of clinical and molecular evidence.