Browsing by Author "Larsson, Olle"

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    Comprehensive Genetic Analysis Identifies a Pathognomonic NAB2/STAT6 Fusion Gene, Nonrandom Secondary Genomic Imbalances, and a Characteristic Gene Expression Profile in Solitary Fibrous Tumor
    (Genes, Chromosomes and Cancer, 2013) Mohajeri, Arezoo; Tayebwa, Johnbosco; Collin, Anna; Nilsson, Jenny; Magnusson, Linda; Vult von Steyern, Fredrik; Brosj, Otte; Domanski, Henryk A.; Larsson, Olle; Sciot, Raf; Debiec-Rychter, Maria; Hornick, Jason L.; Mandahl, Nils; Nord, Karolin H.; Mertens, Fredrik
    Solitary fibrous tumor (SFT) is a mesenchymal neoplasm displaying variable morphologic and clinical features. To identify pathogenetically important genetic rearrangements, 44 SFTs were analyzed using a variety of techniques. Chromosome banding and fluorescence in situ hybridization (FISH) showed recurrent breakpoints in 12q13, clustering near the NAB2 and STAT6 genes, and single nucleotide polymorphism array analysis disclosed frequent deletions affecting STAT6. Quantitative real-time PCR revealed high expression levels of the 5’-end of NAB2 and the 3’-end of STAT6, which at deep sequencing of enriched DNA corresponded to NAB2/STAT6 fusions. Subsequent reverse-transcriptase PCR (RT-PCR) analysis identified a NAB2/STAT6 fusion in 37/41 cases, confirming that this fusion gene underlies the pathogenesis of SFT. The hypothesis that the NAB2/STAT6 fusions will result in altered properties of the transcriptional co-repressor NAB2 - a key regulator of the early growth response 1 (EGR1) transcription factor – was corroborated by global gene expression analysis; SFTs showed deregulated expression of EGR1 target genes, as well as of other, developmentally important genes. We also identified several nonrandom secondary changes, notably loss of material from 13q and 14q. As neither chromosome banding nor FISH analysis identify more than a minor fraction of the fusion-positive cases, and because multiple primer combinations are required to identify all possible fusion transcripts by RT-PCR, alternative diagnostic markers might instead be found among deregulated genes identified at global gene expression analysis. Indeed, using immunohistochemistry on tissue microarrays, the top up-regulated gene, GRIA2, was found to be differentially expressed also at the protein level.
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    Fusions involving protein kinase C and membrane-associated proteinsin benign fibrous histiocytoma
    (he International Journal of Biochemistry & Cell Biology,, 2014) Płaszczyca, Anna; Nilsson, Jenny; Magnusson, Linda; Brosjö, Otte; Larsson, Olle; Vult von Steyernd, Fredrik; Domanski, Henryk A.; Lilljebjörn, Henrik; Fioretos, Thoas; Tayebwa, Johnbosco; Mandahl, Nils; Nord, Karolin H.; Mertens, Fredrik
    Benign fibrous histiocytoma (BFH) is a mesenchymal tumor that most often occurs in the skin (so-called dermatofibroma), but may also appear in soft tissues (so-called deep BFH) and in the skeleton(so-called non-ossifying fibroma). The origin of BFH is unknown, and it has been questioned whether itis a true neoplasm. Chromosome banding, fluorescence in situ hybridization, single nucleotide polymor-phism arrays, RNA sequencing, RT-PCR and quantitative real-time PCR were used to search for re current somatic mutations in a series of BFH. BFHs were found to harbor recurrent fusions of genes encoding membrane-associated proteins (podoplanin, CD63 and LAMTOR1) with genes encoding protein kinaseC (PKC) isoforms PRKCB and PRKCD. PKCs are serine–threonine kinases that through their many phos-phorylation targets are implicated in a variety of cellular processes, as well as tumor development. When inactive, the amino-terminal, regulatory domain of PKCs suppresses the activity of their catalytic domain. Upon activation, which requires several steps, they typically translocate to cell membranes, where they interact with different signaling pathways. The detected PDPN-PRKCB, CD63-PRKCD and LAMTOR1-PRKCDgene fusions are all predicted to result in chimeric proteins consisting of the membrane-binding part of PDPN, CD63 or LAMTOR1 and the entire catalytic domain of the PKC. This novel pathogenetic mechanism should result in constitutive kinase activity at an ectopic location. The results show that BFH indeed is atrue neoplasm, and that distorted PKC activity is essential for tumorigenesis. The findings also provide means to differentiate BFH from other skin and soft tissue tumors.
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    Recurrent PRDM10 Gene Fusions in Undifferentiated Pleomorphic Sarcoma
    (Clinical Cancer Research, 2015) Hofvander, Jakob; Tayebwa, Johnbosco; Nilsson, Jenny; Magnusson, Linda; Brosjo, Otte; Larsson, Olle; Vult von Steyern, Fredrik; Mandahl, Nils; Fletcher, Christopher D.M.; Mertens, Fredrik
    Undifferentiated pleomorphic sarcoma (UPS) is defined as a sarcoma with cellular pleomorphism and no identifiable line of differentiation. It is typically a high-grade lesion with a metastatic rate of about one third. No tumor-specific rearrangement has been identified, and genetic markers that could be used for treatment stratification are lacking. We performed transcriptome sequencing (RNA-Seq) to search for novel gene fusions. Experimental design: RNA-Seq, FISH, and/or various PCR methodologies were used to search for gene fusions and rearrangements of the PRDM10 gene in 84 soft tissue sarcomas. Results: Using RNA-Seq, two cases of UPS were found to display novel gene fusions, both involving the transcription factor PRDM10 as the 30 partner and either MED12 or CITED2 as the 50 partner gene. Further screening of 82 soft tissue sarcomas for rearrangements of the PRDM10 locus revealed one more UPS with a MED12/PRDM10 fusion. None of these genes has been implicated in neoplasia-associated gene fusions before. Conclusions: Our results suggest that PRDM10 fusions are present in around 5% of UPS. Although the fusion-positive cases in our series showed the same nuclear pleomorphism and lack of differentiation as other UPS, it is noteworthy that all three were morphologically low grade and that none of the patients developed metastases. Thus, PRDM10 fusion-positive sarcomas may constitute a clinically important subset of UPS.
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    RNA sequencing of sarcomas with simple karyotypes: identification and enrichment of fusion transcripts
    (Laboratory Investigation, 2015) Hofvander, Jakob; Tayebwa, Johnbosco; Nilsson, Jenny; Magnusson, Linda; Brosjö, Otte; Larsson, Olle; Vult von Steyern, Fredrik; Domanski, Henryk A.; Mandahl, Nils; Mertens, Fredrik
    Gene fusions are neoplasia-associated mutations arising from structural chromosomal rearrangements. They have a strong impact on tumor development and constitute important diagnostic markers. Malignant soft tissue tumors (sarcomas) constitute a heterogeneous group of neoplasms with 450 distinct subtypes, each of which is rare. In addition, there is considerable morphologic overlap between sarcomas and benign lesions. Several subtypes display distinct gene fusions, serving as excellent biomarkers. The development of methods for deep sequencing of the complete transcriptome (RNA-Seq) has substantially improved the possibilities for detecting gene fusions. With the aim of identifying new gene fusions of biological and clinical relevance, eight sarcomas with simple karyotypes, ie, only one or a few structural rearrangements, were subjected to massively parallel paired-end sequencing of mRNA. Three different algorithms were used to identify fusion transcripts from RNA-Seq data. Three novel (KIAA2026-NUDT11, CCBL1-ARL1, and AFF3-PHF1) and two previously known fusions (FUS-CREB3L2 and HAS2-PLAG1) were found and could be verified by other methods. These findings show that RNA-Seq is a powerful tool for detecting gene fusions in sarcomas but also suggest that it is advisable to use more than one algorithm to analyze the output data as only two of the confirmed fusions were reported by more than one of the gene fusion detection software programs. For all of the confirmed gene fusions, at least one of the genes mapped to a chromosome band implicated by the karyotype, suggesting that sarcomas with simple karyotypes constitute an excellent resource for identifying novel gene fusions.