Signatures of copy number alterations in human cancer.

Gains and losses of DNA are prevalent in cancer and emerge as a consequence of inter-related processes of replication stress, mitotic errors, spindle multipolarity and breakage-fusion-bridge cycles, among others, which may lead to chromosomal instability and aneuploidy1,2. These copy number alterations contribute to cancer initiation, progression and therapeutic resistance3-5. Here we present a conceptual framework to examine the patterns of copy number alterations in human cancer that is widely applicable to diverse data types, including whole-genome sequencing, whole-exome sequencing, reduced representation bisulfite sequencing, single-cell DNA sequencing and SNP6 microarray data. Deploying this framework to 9,873 cancers representing 33 human cancer types from The Cancer Genome Atlas6 revealed a set of 21 copy number signatures that explain the copy number patterns of 97% of samples. Seventeen copy number signatures were attributed to biological phenomena of whole-genome doubling, aneuploidy, loss of heterozygosity, homologous recombination deficiency, chromothripsis and haploidization. The aetiologies of four copy number signatures remain unexplained. Some cancer types harbour amplicon signatures associated with extrachromosomal DNA, disease-specific survival and proto-oncogene gains such as MDM2. In contrast to base-scale mutational signatures, no copy number signature was associated with many known exogenous cancer risk factors. Our results synthesize the global landscape of copy number alterations in human cancer by revealing a diversity of mutational processes that give rise to these alterations.

Gene amplification in neoplasia: A cytogenetic survey of 80 131 cases.

Gene amplification is a crucial process in cancer development, leading to the overexpression of oncogenes. It manifests cytogenetically as extrachromosomal double minutes (dmin), homogeneously staining regions (hsr), or ring chromosomes (r). This study investigates the prevalence and distribution of these amplification markers in a survey of 80 131 neoplasms spanning hematologic disorders, and benign and malignant solid tumors. The study reveals distinct variations in the frequency of dmin, hsr, and r among different tumor types. Rings were the most common (3.4%) sign of amplification, followed by dmin (1.3%), and hsr (0.8%). Rings were particularly frequent in malignant mesenchymal tumors, especially liposarcomas (47.5%) and osteosarcomas (23.4%), dmin were prevalent in neuroblastoma (30.9%) and pancreatic carcinoma (21.9%), and hsr frequencies were highest in head and neck carcinoma (14.0%) and neuroblastoma (9.0%). Combining all three amplification markers (dmin/hsr/r), malignant solid tumors consistently exhibited higher frequencies than hematologic disorders and benign solid tumors. The structural characteristics of these amplification markers and their potential role in tumorigenesis and tumor progression highlight the complex interplay between cancer-initiating gene-level alterations, for example, fusion genes, and subsequent amplification dynamics. Further research integrating cytogenetic and molecular approaches is warranted to better understand the underlying mechanisms of these amplifications, in particular, the enigmatic question of why certain malignancies display certain types of amplification. Comparing the present results with molecular genetic data proved challenging because of the diversity in definitions of amplification across studies. This study underscores the need for standardized definitions in future work.

A cryptic EWSR1::DDIT3 fusion in myxoid liposarcoma: Potential pitfalls with FISH and cytogenetics.

Myxoid liposarcoma (MLS) is molecularly characterized by fusions involving the DDIT3 gene in chromosome band 12q13; the fusion partner is FUS in band 16p11 in 90-95% of the cases and EWSR1 in band 22q12 in the remaining 5-10%. Hence, molecular studies, often fluorescence in situ hybridization (FISH) for DDIT3 rearrangement, are useful for establishing a correct diagnosis. Although all MLS tumors should have DDIT3 fusions, it is important to be aware of reasons for potential false-negative results. We here present a case of MLS that was negative for FISH for DDIT3, that showed an unexpected t(11;22) at G-banding, but that displayed a characteristic EWSR1::DDIT3 fusion at RNA-sequencing. The results suggest that neoplasia-associated fusions that, due to the transcriptional orientations of the two genes involved, cannot arise through only two double-strand breaks are more likely to be associated with negative FISH-findings and unexpected karyotypes.

Insertion of the CXXC domain of KMT2A into YAP1: An unusual mechanism behind the formation of a chimeric oncogenic protein.

Most neoplasia-associated gene fusions are formed through the fusion of the 5'-part of one gene with the 3'-part of another. We here describe a unique mechanism, by which a part of the KMT2A gene through an insertion replaces part of the YAP1 gene. The resulting YAP1::KMT2A::YAP1 (YKY) fusion was verified by RT-PCR in three cases of sarcoma morphologically resembling sclerosing epithelioid fibrosarcoma (SEF-like sarcoma). In all cases, a portion (exons 4/5-6) encoding the CXXC domain of KMT2A was inserted between exon 4/5 and exon 8/9 of YAP1. The inserted sequence from KMT2A thus replaced exons 5/6-8 of YAP1, which encode an important regulatory sequence of YAP1. To evaluate the cellular impact of the YKY fusion, global gene expression profiles from fresh frozen and formalin-fixed YKY-expressing sarcomas were compared with control tumors. The effects of the YKY fusion, as well as YAP1::KMT2A and KMT2A::YAP1 fusion constructs, were further studied in immortalized fibroblasts. Analysis of differentially upregulated genes revealed significant overlap between tumors and cell lines expressing YKY, as well as with previously reported YAP1 fusions. Pathway analysis of upregulated genes in cells and tumors expressing YKY revealed an enrichment of genes included in key oncogenic signaling pathways, such as Wnt and Hedgehog. As these pathways are known to interact with YAP1, it seems likely that the pathogenesis of sarcomas with the YKY fusion is linked to distorted YAP1 signaling.

A cloud-based resource for genome coordinate-based exploration and large-scale analysis of chromosome aberrations and gene fusions in cancer.

Cytogenetic analysis provides important information on the genetic mechanisms of cancer. The Mitelman Database of Chromosome Aberrations and Gene Fusions in Cancer (Mitelman DB) is the largest catalog of acquired chromosome aberrations, presently comprising >70 000 cases across multiple cancer types. Although this resource has enabled the identification of chromosome abnormalities leading to specific cancers and cancer mechanisms, a large-scale, systematic analysis of these aberrations and their downstream implications has been difficult due to the lack of a standard, automated mapping from aberrations to genomic coordinates. We previously introduced CytoConverter as a tool that automates such conversions. CytoConverter has now been updated with improved interpretation of karyotypes and has been integrated with the Mitelman DB, providing a comprehensive mapping of the 70 000+ cases to genomic coordinates, as well as visualization of the frequencies of chromosomal gains and losses. Importantly, all CytoConverter-generated genomic coordinates are publicly available in Google BigQuery, a cloud-based data warehouse, facilitating data exploration and integration with other datasets hosted by the Institute for Systems Biology Cancer Gateway in the Cloud (ISB-CGC) Resource. We demonstrate the use of BigQuery for integrative analysis of Mitelman DB with other cancer datasets, including a comparison of the frequency of imbalances identified in Mitelman DB cases with those found in The Cancer Genome Atlas (TCGA) copy number datasets. This solution provides opportunities to leverage the power of cloud computing for low-cost, scalable, and integrated analysis of chromosome aberrations and gene fusions in cancer.

Tetraploidization Increases the Motility and Invasiveness of Cancer Cells.

Polyploidy and metastasis are associated with a low probability of disease-free survival in cancer patients. Polyploid cells are known to facilitate tumorigenesis. However, few data associate polyploidization with metastasis. Here, by generating and using diploid (2n) and tetraploid (4n) clones from malignant fibrous histiocytoma (MFH) and colon carcinoma (RKO), we demonstrate the migration and invasion advantage of tetraploid cells in vitro using several assays, including the wound healing, the OrisTM two-dimensional cell migration, single-cell migration tracking by video microscopy, the Boyden chamber, and the xCELLigence RTCA real-time cell migration. Motility advantage was observed despite tetraploid cell proliferation weakness. We could also demonstrate preferential metastatic potential in vivo for the tetraploid clone using the tail vein injection in mice and tracking metastatic tumors in the lung. Using the Mitelman Database of Chromosome Aberrations in Cancer, we found an accumulation of polyploid karyotypes in metastatic tumors compared to primary ones. This work reveals the clinical relevance of the polyploid subpopulation and the strategic need to highlight polyploidy in preclinical studies as a therapeutic target for metastasis.

Amplification of ERBB2 (HER2) in embryonal rhabdomyosarcoma: A potential treatment target in rare cases?

The ERBB2 gene encodes a receptor tyrosine kinase also known as HER2. The gene is amplified and overexpressed in one-fifth of breast carcinomas; patients with such tumors benefit from targeted treatment with trastuzumab or other drugs blocking the receptor. In addition, ERBB2 has been shown to be amplified and/or overexpressed in a variety of other malignancies. Notably, both alveolar and embryonal rhabdomyosarcoma (RMS), especially in children, often show increased expression of ERBB2. Although high-level amplification of the gene has not been described in RMS, its frequent expression at the cell surface of RMS cells has been exploited for chimeric antigen receptor T-cell (CAR T)-based treatment strategies. We here describe two cases of pediatric, fusion-negative embryonal RMS with high-level amplification of the ERBB2 gene. One patient is currently treated with conventional chemotherapy for a recently detected standard risk RMS, whereas the other patient died from metastatic disease. Both tumors displayed focal amplicons (210 and 274 Kb, respectively) in chromosome band 17q12, with proximal and distal borders corresponding to those typically seen in breast cancer. In both tumors, the ERBB2 amplicon correlated with high expression at the RNA and protein levels. Thus, breast cancer-like ERBB2 amplification is a very rare, but recurrent feature of pediatric RMS, and should be exploited as an alternative treatment target.

The diagnostic utility of DNA copy number analysis of core needle biopsies from soft tissue and bone tumors.

Morphologic and immunohistochemical analysis of preoperative core needle biopsies (CNB) is important in the management of patients with soft tissue and bone tumors (STBTs). Most SBTB subtypes have more or less extensive DNA copy number aberrations (CNA), potentially providing useful diagnostic information. To evaluate the technical feasibility of single nucleotide polymorphism (SNP) array analysis and the diagnostic usefulness of the copy number profiles, we studied CNBs from 171 patients with suspected STBTs. SNP array analysis could be performed on 168 (98%) of the samples. The CNA profile was compatible with the CNB diagnosis in 87% of the cases. Discrepant cases were dominated by false-negative results due to nonrepresentative material or contamination with normal cells. 70 genomic profiles were indicative of specific histopathologic tumor entities and in agreement with the corresponding CNB diagnoses in 83%. In 96 of the cases with aberrant CNA profiles, the SNP profiles were of sufficient quality for segmentation, allowing clustering analysis on the basis of the Jaccard similarity index. The analysis of these segment files showed three major CNA clusters, based on the complexity levels and the predominance of gains versus losses. For 43 of these CNB samples, we had SNP array data also from their corresponding surgical samples. In 33 of these pairs, the two corresponding samples clustered next to each other, with Jaccard scores ranging from 0.61 to 0.99 (median 0.96). Also, for those tumor pairs that did not cluster together, the Jaccard scores were relatively high (median 0.9). 10 cases showed discrepant results, mainly due to varying degrees of normal cell contamination or technical issues. Thus, the copy number profile seen in a CNB is typically highly representative of the major cell population in the tumor.

Overlapping morphological, immunohistochemical and genetic features of superficial CD34-positive fibroblastic tumor and PRDM10-rearranged soft tissue tumor.

Superficial CD34-positive fibroblastic tumor (SCD34FT) is a recently recognized soft tissue tumor that is considered to be of borderline malignancy. The pathogenesis of this tumor remains incompletely understood, but it has been suggested that SCD34FT overlaps with tumors showing fusions involving the PRDM10 gene. Previous analyses of PRDM10-rearranged tumors have demonstrated that they have a distinct gene expression profile, resulting in high expression of CADM3 (also known as SynCam3), which can be detected immunohistochemically. Here, we investigated a series (n = 43) of SCD34FT or PRDM10-rearranged tumors and potential mimics (n = 226) with regard to morphological, genetic, and immunohistochemical features. The results show that SCD34FT and PRDM10-rearranged tumor are morphologically indistinguishable; 41 of 43 tumors of both entities are CADM3-positive. Hence, we suggest that they constitute a single entity, preferably referred to as SCD34FT. Expression of CADM3 was only rarely seen in other soft tissue tumors, except in tumors with Schwann cell differentiation. Thus, IHC for CADM3, in combination with the characteristic morphological features, is a valuable adjunct in the diagnosis of SCD34FT.

Genomic and transcriptomic characterization of desmoplastic small round cell tumors.

Desmoplastic small round cell tumor (DSRCT) is a highly aggressive soft tissue tumor primarily affecting children and young adults. Most cases display a pathognomonic EWSR1-WT1 gene fusion, presumably constituting the primary driver event. Little is, however, known about secondary genetic changes that may affect tumor progression. We here studied 25 samples from 19 DSRCT patients using single nucleotide polymorphism arrays and found that all samples had copy number alterations. The most common imbalances were gain of chromosomes/chromosome arms 1/1q and 5/5p and loss of 6/6q and 16/16q, all occurring in at least eight of the patients. Five cases showed homozygous deletions, affecting a variety of known tumor suppressor genes, for example, CDKN2A and NF1. As almost all patients died of their disease, the impact of individual imbalances on survival could not be evaluated. Global gene expression analysis using mRNA sequencing on fresh-frozen samples from seven patients revealed a distinct transcriptomic profile, with enrichment of genes involved in neural differentiation. Two genes - GJB2 and GAL - that showed higher expression in DSRCT compared to control tumors could be further investigated for their potential as diagnostic markers at the protein level.

"Inflammatory Leiomyosarcoma" and "Histiocyte-rich Rhabdomyoblastic Tumor": a clinicopathological, immunohistochemical and genetic study of 13 cases, with a proposal for reclassification as "Inflammatory Rhabdomyoblastic Tumor".

Inflammatory leiomyosarcoma (ILMS), defined as "a malignant neoplasm showing smooth muscle differentiation, a prominent inflammatory infiltrate, and near-haploidization", is a very rare soft tissue tumor with a generally favorable prognosis. The morphologic features of "histiocyte-rich rhabdomyoblastic tumor" (HRRMT) are similar to those of ILMS, although this lesion shows by definition a skeletal muscle phenotype. Recent gene expression profiling and immunohistochemical studies have also suggested that ILMS and HRRMT may be related. We studied the clinicopathologic, immunohistochemical and genetic features of four cases previously classified as ILMS and nine classified as HRRMT. Tumors from both groups tended to occur in the deep soft tissues of the extremities of young to middle-aged males and exhibited indolent behavior. Morphologically, all were well-circumscribed, often encapsulated, and showed a striking histiocyte-rich inflammatory infiltrate admixed with variably pleomorphic tumor cells showing spindled and epithelioid to rhabdoid morphology, eosinophilic cytoplasm, and prominent nucleoli, but few, if any, mitotic figures. Immunohistochemically, the tumor cells expressed desmin, alpha-smooth muscle actin, and the rhabdomyoblastic markers PAX7, MyoD1, and myogenin. H-caldesmon expression was absent in all cases, using the specific h-CD antibody. Karyotypic study (1 HRRMT) and genome-wide copy number analysis (7 HRRMT, OncoScan SNP assay), revealed near-haploidization in four cases, with subsequent genome doubling in one, an identical phenotype to that seen in ILMS. We propose reclassification of ILMS and HRRMT as "inflammatory rhabdomyoblastic tumor", a name which accurately describes the salient morphologic and immunohistochemical features of this distinctive tumor, as well as its intermediate (rarely metastasizing) clinical behavior.

Deep sequencing of myxoinflammatory fibroblastic sarcoma.

Myxoinflammatory fibroblastic sarcoma (MIFS) has recurrent genetic features in the form of a translocation t(1;10)(p22-31;q24-25), BRAF gene fusions, and/or an amplicon in 3p11-12 including the VGLL3 gene. The breakpoints on chromosomes 1 and 10 in the t(1;10) cluster in or near the TGFBR3 and OGA genes, respectively. We here used a combination of deep sequencing of the genome (WGS), captured sequences (Cap-seq), and transcriptome (RNA-seq) and genomic arrays to investigate the molecular outcome of the t(1;10) and the VGLL3 amplicon, as well as to assess the spectrum of other recurrent genomic features in MIFS. Apart from a ROBO1-BRAF chimera in a t(1;10)-negative MIFS-like tumor, no fusion gene was found at RNA-seq. This was in line with WGS and Cap-seq results, revealing variable breakpoints in chromosomes 1 and 10 and genomic breakpoints that should not yield functional fusion transcripts. The most common genomic rearrangements were breakpoints in or around the OGA, NPM3, and FGF8 genes in chromosome band 10q24, and loss of 1p11-p21 and 10q26-qter (all simultaneously present in 6/7 MIFS); a breakpoint in or near TGFBR3 in chromosome 1 was found in four of these tumors. Amplification and overexpression of VGLL3 was a consistent feature in MIFS and MIFS-like tumors with amplicons in 3p11-12. The significant molecular genetic outcome of the recurrent t(1;10) could be loss of genetic material from 1p and 10q. Other recurrent genomic imbalances in MIFS, such as homozygous loss of CDKN2A and 3p- and 13q-deletions, are shared with other sarcomas, suggesting overlapping pathogenetic pathways.

PHF1 fusions cause distinct gene expression and chromatin accessibility profiles in ossifying fibromyxoid tumors and mesenchymal cells.

Ossifying fibromyxoid tumor (OFMT) is a soft tissue tumor frequently displaying gene fusions, most of which affect the PHF1 gene. PHF1 encodes plant homeodomain finger protein 1, which is involved in various processes regulating gene transcription, including those orchestrated by the polycomb repressor complex 2. Here, a series of 37 OFMTs, including 18 typical, 9 atypical, and 10 malignant variants, was analyzed with regard to transcriptomic features, gene fusion and copy number status, and/or single-nucleotide variants. The effects on gene expression and chromatin accessibility of three detected fusions (EP400-PHF1, MEAF6-PHF1, and PHF1-TFE3) were further evaluated in fibroblasts. Genomic imbalances showed a progression-related pattern, with more extensive copy number changes among atypical/malignant lesions than among typical OFMTs; loss of the RB1 gene was restricted to atypical/malignant OFMTs, occurring in one-third of the cases. RNA sequencing identified fusion transcripts in >80% of the cases analyzed, including a novel CSMD1-MEAF6. The gene-expression profile of OFMT was distinct from that of other soft tissue tumors, with extensive transcriptional upregulation of genes in OFMT. These findings were largely recapitulated in gene fusion-expressing fibroblast lines, suggesting that genes involved in, e.g., Wnt signaling and/or being regulated through trimethylation of lysine 27 in histone 3 (H3K27me3) are pivotal for OFMT development. The genes showing differentially higher expression in fusion-expressing cells paralleled increased chromatin accessibility, as revealed by ATAC sequencing. Thus, the present study suggests that OFMT develops through gene fusions that have extensive epigenetic consequences.

Undifferentiated pleomorphic sarcomas with PRDM10 fusions have a distinct gene expression profile.

Undifferentiated pleomorphic sarcoma (UPS) is a highly aggressive soft tissue tumor. A subset of UPS is characterized by a CITED2-PRDM10 or a MED12-PRDM10 gene fusion. Preliminary data suggest that these so-called PRDM10-rearranged tumors (PRT) are clinically more indolent than classical high-grade UPS, and hence important to recognize. Here, we assessed the spectrum of accompanying mutations and the gene expression profile in PRT using genomic arrays and sequencing of the genome (WGS) and transcriptome (RNA-seq). The fusion protein's function was further investigated by conditional expression of the CITED2-PRDM10 fusion in a fibroblast cell line, followed by RNA-seq and an assay for transposase-accessible chromatin (ATAC-seq). The CADM3 gene was found to be differentially up-regulated in PRT and cell lines and was also evaluated for expression at the protein level using immunohistochemistry (IHC). The genomic analyses identified few and nonrecurrent mutations in addition to the structural variants giving rise to the gene fusions, strongly indicating that the PRDM10-fusions represent the critical driver mutations. RNA-seq of tumors showed a distinct gene expression profile, separating PRT from high-grade UPS and other soft tissue tumors. CADM3 was among the genes that was consistently and highly expressed in both PRT and fibroblasts expressing CITED2-PRDM10, suggesting that it is a direct target of the PRDM10 transcription factor. This conclusion is in line with sequencing data from ATAC-seq, showing enrichment of PRDM10 binding sites, suggesting that the amino-terminal fusion partner contributes by making the DNA more accessible to PRDM10 binding. © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Cancer chromosome breakpoints cluster in gene-rich genomic regions.

Cancer cells are characterized by chromosome abnormalities, of which some, in particular balanced rearrangements, are associated with distinct tumor entities and/or with specific gene rearrangements that represent important steps in the carcinogenic process. However, the vast majority of cytogenetically detectable structural aberrations in cancer cells have not been characterized at the nucleotide level; hence, their importance and functional consequences are unknown. By ascertaining the chromosomal breakpoints in 22 344 different clonal structural chromosome abnormalities identified in the karyotypes of 49 626 cases of neoplastic disorders we here show that the distribution of breakpoints is strongly associated (P < 0.0001) with gene content within the affected chromosomal bands. This association also remains highly significant in separate analyses of recurrent and nonrecurrent chromosome abnormalities as well as of specific subtypes of cancer (P < 0.0001 for all comparisons). In contrast, the impact of band length was negligible. The breakpoint distribution is thus not stochastic-gene-rich regions are preferentially affected. Several genomic features relating to transcription, replication, and chromatin organization have been found to enhance chromosome breakage frequencies; this indicates that gene-rich regions may be more break-prone. The salient finding in the present study is that a substantial fraction of all structural chromosome abnormalities, not only those specifically associated with certain tumor types, may affect genes that are pathogenetically important. If this interpretation is correct, then the prevailing view that the great majority of cancer chromosome aberrations is cytogenetic noise can be seriously questioned.

Most gene fusions in cancer are stochastic events.

Cancer-associated gene fusions resulting in chimeric proteins or aberrant expression of one or both partner genes are pathogenetically and clinically important in several hematologic malignancies and solid tumors. Since the advent of different types of massively parallel sequencing (MPS), the number of identified gene fusions has increased dramatically, prompting the question whether they all have a biologic impact. By ascertaining the chromosomal locations of 8934 genes involved in 10 861 gene fusions reported in the literature, we here show that there is a highly significant association between gene content of chromosomes and chromosome bands and number of genes involved in fusions. This strongly suggests that a clear majority of gene fusions detected by MPS are stochastic events associated with the number of genes available to participate in fusions and that most reported gene fusions are passengers without any pathogenetic importance.

Aberrant receptor tyrosine kinase signaling in lipofibromatosis: a clinicopathological and molecular genetic study of 20 cases.

Lipofibromatosis is a rare pediatric soft tissue tumor with predilection for the hands and feet. Previously considered to represent "infantile fibromatosis", lipofibromatosis has distinctive morphological features, with mature adipose tissue, short fascicles of bland fibroblastic cells, and lipoblast-like cells. Very little is known about the genetic underpinnings of lipofibromatosis. Prompted by our finding of the FN1-EGF gene fusion, previously shown to be a characteristic feature of calcifying aponeurotic fibroma (CAF), in a morphologically typical case of lipofibromatosis that recurred showing features of CAF, we studied a cohort of 20 cases of lipofibromatosis for this and other genetic events. The cohort was composed of 14 males and 6 females (median age 3 years; range 1 month-14 years). All primary tumors showed classical lipofibromatosis morphology. Follow-up disclosed three local recurrences, two of which contained calcifying aponeurotic fibroma-like nodular calcifications in addition to areas of classic lipofibromatosis, and no metastases. By FISH and RNA sequencing, four cases were positive for FN1-EGF and one case each showed an EGR1-GRIA1, TPR-ROS1, SPARC-PDGFRB, FN1-TGFA, EGFR-BRAF, VCL-RET, or HBEGF-RBM27 fusion. FN1-EGF was the only recurrent fusion, suggesting that some cases of "lipofibromatosis" may represent calcifying aponeurotic fibroma lacking hallmark calcifications. Several of the genes involved in fusions (BRAF, EGFR, PDGFRB, RET, and ROS1) encode receptor tyrosine kinases (RTK), or ligands to the RTK EGFR (EGF, HBEGF, TGFA), suggesting a shared deregulation of the PI3K-AKT-mTOR pathway in a large subset of lipofibromatosis cases.

Gene fusion involving the insulin-like growth factor 1 receptor in an ALK-negative inflammatory myofibroblastic tumour.

AIMS: Inflammatory myofibroblastic tumour (IMT) is a soft tissue tumour primarily affecting children and young adults. Approximately 50% of IMTs have gene fusions involving the receptor tyrosine kinase (RTK)-encoding ALK gene, providing a molecular rationale for treating IMT patients with unresectable tumours with tyrosine kinase inhibitors (TKI). However, a subset of IMT instead displays fusions affecting other RTKencoding genes, so far including NTRK3, PDGFRB and ROS1. Also, IMTs with variant RTK fusions may respond well to TKI treatment, but can be dif?cult to identify as they are negative for ALK staining at immunohistochemistry, the standard method for detection of ALK rearrangements. MATERIALS AND METHODS: We used RNA-sequencing to search for alternate fusion events in an ALK-negative IMT. RESULTS AND CONCLUSIONS: We found a novel fusion gene - FN1-IGF1R. The FN1 gene, encoding ?bronectin, is thought to provide a strong promoter activity for the kinase domain of the RTK insulin-like growth factor 1 receptor, a mechanism similar to previously described RTK fusions in IMT.

Inflammatory leiomyosarcoma is a distinct tumor characterized by near-haploidization, few somatic mutations, and a primitive myogenic gene expression signature.

Inflammatory leiomyosarcoma is a soft-tissue tumor resembling conventional leiomyosarcoma, but with a prominent intrinsic inflammatory component. Previous studies have suggested that inflammatory leiomyosarcoma differs genetically from leiomyosarcoma, but in-depth analyses are lacking. Here we provide a comprehensive picture of the genome and transcriptome of inflammatory leiomyosarcoma by combining cytogenetic, single-nucleotide polymorphism array, mRNA-sequencing, and whole-exome sequencing data. The results show that inflammatory leiomyosarcoma has a specific genetic profile characterized by near-haploidization with or without subsequent whole-genome doubling. Consistently, both parental copies of chromosomes 5 and 22 are preserved. Apart from recurrent mutation of the NF1 gene, additional somatic events that could serve as driver mutations were not found at either the nucleotide or the genome level. Furthermore, no fusion transcripts were identified. Global gene expression profiling revealed particularly prominent differential expression of genes, including ITGA7, MYF5, MYF6, MYOD1, MYOG, and PAX7, involved in muscle development and function, providing strong argument for grouping inflammatory leiomyosarcoma with myogenic sarcomas, rather than with myofibroblastic lesions. Combined with previously published data, there are now 10 cases of inflammatory leiomyosarcoma with confirmed near-haploid genotype. These patients differ from leiomyosarcoma patients in being younger (median 41 years), showing a male predominance (9:1), and few relapses (1 of 8 informative patients). Thus, the clinical, morphological, and genetic data provide compelling support for inflammatory leiomyosarcoma being a distinct subtype of myogenic tumors.

Inferior survival for patients with malignant peripheral nerve sheath tumors defined by aberrant TP53.

Malignant peripheral nerve sheath tumor is a rare and aggressive disease with poor treatment response, mainly affecting adolescents and young adults. Few molecular biomarkers are used in the management of this cancer type, and although TP53 is one of few recurrently mutated genes in malignant peripheral nerve sheath tumor, the mutation prevalence and the corresponding clinical value of the TP53 network remains unsettled. We present a multi-level molecular study focused on aberrations in the TP53 network in relation to patient outcome in a series of malignant peripheral nerve sheath tumors from 100 patients and 38 neurofibromas, including TP53 sequencing, high-resolution copy number analyses of TP53 and MDM2, and gene expression profiling. Point mutations in TP53 were accompanied by loss of heterozygosity, resulting in complete loss of protein function in 8.2% of the malignant peripheral nerve sheath tumors. Another 5.5% had MDM2 amplification. TP53 mutation and MDM2 amplification were mutually exclusive and patients with either type of aberration in their tumor had a worse prognosis, compared to those without (hazard ratio for 5-year disease-specific survival 3.5, 95% confidence interval 1.78-6.98). Both aberrations had similar consequences on the gene expression level, as analyzed by a TP53-associated gene signature, a property also shared with the copy number aberrations and/or loss of heterozygosity at the TP53 locus, suggesting a common "TP53-mutated phenotype" in as many as 60% of the tumors. This was a poor prognostic phenotype (hazard ratio = 4.1, confidence interval:1.7-9.8), thus revealing a TP53-non-aberrant patient subgroup with a favorable outcome. The frequency of the "TP53-mutated phenotype" warrants explorative studies of stratified treatment strategies in malignant peripheral nerve sheath tumor.