Identification of novel GNAS mutations in intramuscular myxoma using next-generation sequencing with single-molecule tagged molecular inversion probes.

BACKGROUND: Intramuscular myxoma (IM) is a hypocellular benign soft tissue neoplasm characterized by abundant myxoid stroma and occasional hypercellular areas. These tumors can, especially on biopsy material, be difficult to distinguish from low-grade fibromyxoid sarcoma or low-grade myxofibrosarcoma. GNAS mutations are frequently involved in IM, in contrast to these other malignant tumors. Therefore, sensitive molecular techniques for detection of GNAS aberrations in IM, which frequently yield low amounts of DNA due to poor cellularity, will be beneficial for differential diagnosis. METHODS: In our study, a total of 34 IM samples from 33 patients were analyzed for the presence of GNAS mutations, of which 29 samples were analyzed using a gene-specific TaqMan genotyping assay for the detection of GNAS hotspot mutations c.601C > T and c602G > A in IM, and 32 samples using a novel next generation sequencing (NGS)-based approach employing single-molecule tagged molecular inversion probes (smMIP) to identify mutations in exon 8 and 9 of GNAS. Results between the two assays were compared for their ability to detect GNAS mutations with high confidence. RESULTS: In total, 23 of 34 samples were successfully analyzed with both techniques showing GNAS mutations in 12 out of 23 (52%) samples. The remaining 11 samples were analyzed with either TaqMan assay or smMIP assay only. The TaqMan assay revealed GNAS mutations in 16 out of 29 samples (55%), with six samples c.601C > T (p.R201C; 38%) and ten samples c.602G > A (p.R201H; 62%) missense mutations. The smMIP assay identified mutations in 16 out of 28 samples (57%), with five samples c.601C > T (p.R201C; 31%) and seven samples c.602G > A (p.R201H; 44%) missense mutations. In addition, four samples (25%) revealed novel IM-associated mutations, including c.601C > A (p.R201S), c.602G > T (p.R201L), c.602G > C (p.R201P) and c.680A > G (p.Q227R). Combining the results of both tests, 23 out of 34 sporadic IM samples (68%) showed a GNAS mutation. CONCLUSIONS: Both the TaqMan and the smMIP assay a show a high degree of concordance in detecting GNAS hotspot mutations in IM with comparable sensitivity. However, since the NGS-based smMIP assay permits mutation detection in whole exons of GNAS, a broader range of GNAS mutations can be identified by the smMIP approach.

Solitary fibrous tumor - clinicopathologic, immunohistochemical and molecular analysis of 28 cases.

BACKGROUND: Solitary fibrous tumor is a mesenchymal tumor of fibroblastic type, which can affect any region of the body. Recently, a recurrent gene fusion NAB2-STAT6 has been identified as molecular hallmark. The NAB2-STAT6 fusion leads to EGR1 activation and transcriptional deregulation of EGR1-dependent target genes and is a driving event in initiation of SFT. In this study, we report the clinicopathologic and RT-PCR findings and evaluated expression of STAT6 and EGR1 protein in a cohort of 28 SFTs. METHODS: 28 patients with a median age of 54 years were included with SFTs originating at different sites, most occurring in the lung and pleura (9, 32%), 5 in soft tissues of the lower extremities (18%) and 5 in the head and neck (18%). For detection of the NAB2-STAT6 fusion gene, RT-PCR was performed using RNA extracted from formalin-fixed and paraffin-embedded tissues. Immunohistochemistry was performed on all cases with antibodies against STAT6 and EGR1. RESULTS: All patients were treated by surgery, 3 with adjuvant chemo- or radiotherapy. Follow-up data of 18 patients could be obtained of which 2 patients died of metastatic disease 13 months and 52 years after first diagnosis. Sixteen patients have no evidence of disease with a median follow up of 29.5 months (range 7 - 120 months). NAB2-STAT6 fusion transcripts were found in 19/28 cases (68%). The most common fusion was between NAB2 exon 4 and STAT6 exon 3 (11/19, 58%), mainly occurring in pleuropulmonary lesions. All cases showed strong nuclear expression of STAT6 (28/28, 100%) while EGR1 showed low-level variable nuclear expression in all samples, comparable with the EGR1 expression results of the control group. CONCLUSIONS: The identification of the NAB2-STAT6 fusion in SFTs can provide important diagnostic information, especially in cases with aberrant morphology or when biopsy material is limited. STAT6 immunohistochemistry is another useful tool in diagnosing SFT. EGR1 immunohistochemistry indicates low-level protein expression in accordance with EGR1 activation due to distorted NAB2 activity. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/13000_2014_224.

[A woman with ongoing anaemia]. / Een vrouw met persisterende anemie.

In a 65-year-old female with iron deficiency anaemia, video capsule endoscopy showed an ulcerative polyp in the proximal ileum. After histological examination we made the diagnosis 'granuloma pyogenicum', a rare cause of gastrointestinal bleeding. After partial resection of the small intestine, the anaemia was cured.

Oestrogen receptor-beta as a potential target for treatment in advanced colorectal cancer: a pilot study.

AIMS: Oestrogen receptor-beta (ER-ß) is expressed in colorectal cancer. Theoretically, ER-ß stimulation could slow down tumour proliferation, and this is supported by preclinical research data. While preparing a Phase II trial for advanced colorectal cancer patients we performed a pilot study with three questions: (i) in what percentage of patients do metastases display strong ER-ß1 expression; (ii) is there any concordance in expression between primary tumours and metastases; and (iii) is the immunohistochemical (IHC) scoring procedure reproducible? METHODS AND RESULTS: Thirty patients were selected, 15 with locoregional lymph node metastases and 15 with either synchronous or metachronous hepatic metastases. All primary tumours and metastases were analysed for immunohistochemical ER-ß1 expression according to a predefined scoring system. The scoring was performed independently by two pathologists in order to calculate the weighted kappa value. Strong ER-ß1 expression was found in four of 15 hepatic metastases and four of 15 lymph node metastases. In 15 of 30 patients, the level of ER-ß1 expression in the metastasis was concordant with that observed in the primary tumour. Weighted kappa values of IHC ER-ß1 expression were satisfactory. CONCLUSIONS: In twenty-five per cent of patients there was strong ER-ß1 expression in metastases, biopsy of which will be considered mandatory for trial inclusion.