Identification of a Novel Oncogenic Fusion Gene SPON1-TRIM29 in Clinical Ovarian Cancer That Promotes Cell and Tumor Growth and Enhances Chemoresistance in A2780 Cells

Gene structure alterations, such as chromosomal rearrangements that develop fusion genes, often contribute to tumorigenesis. It has been shown that the fusion genes identified in public RNA-sequencing datasets are mainly derived from intrachromosomal rearrangements. In this study, we explored fusion transcripts in clinical ovarian cancer specimens based on our RNA-sequencing data. We successfully identified an in-frame fusion transcript SPON1-TRIM29 in chromosome 11 from a recurrent tumor specimen of high-grade serous carcinoma (HGSC), which was not detected in the corresponding primary carcinoma, and validated the expression of the identical fusion transcript in another tumor from a distinct HGSC patient. Ovarian cancer A2780 cells stably expressing SPON1-TRIM29 exhibited an increase in cell growth, whereas a decrease in apoptosis was observed, even in the presence of anticancer drugs. The siRNA-mediated silencing of SPON1-TRIM29 fusion transcript substantially impaired the enhanced growth of A2780 cells expressing the chimeric gene treated with anticancer drugs. Moreover, a subcutaneous xenograft model using athymic mice indicated that SPON1-TRIM29-expressing A2780 cells rapidly generated tumors in vivo compared to control cells, whose growth was significantly repressed by the fusion-specific siRNA administration. Overall, the SPON1-TRIM29 fusion gene could be involved in carcinogenesis and chemotherapy resistance in ovarian cancer, and offers potential use as a diagnostic and therapeutic target for the disease with the fusion transcript.

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Transcriptome Analysis Reveals MFGE8-HAPLN3 Fusion as a Novel Biomarker in Triple-Negative Breast Cancer

Frontiers in Oncology ◽

10.3389/fonc.2021.682021 ◽

2021 ◽

Vol 11 ◽

Author(s):

Meng-Yuan Wang ◽

Man Huang ◽

Chao-Yi Wang ◽

Xiao-Ying Tang ◽

Jian-Gen Wang ◽

...

Keyword(s):

Breast Cancer ◽

Rna Sequencing ◽

Triple Negative Breast Cancer ◽

Triple Negative ◽

Fusion Transcript ◽

Sequencing Data ◽

Chromosome 11 ◽

Fusion Transcripts ◽

Tumor Tissues ◽

Fusion Type

BackgroundTriple-negative breast cancer (TNBC) is a highly aggressive cancer with poor prognosis. The lack of effective targeted therapies for TNBC remains a profound clinical challenge. Fusion transcripts play critical roles in carcinogenesis and serve as valuable diagnostic and therapeutic targets in cancer. The present study aimed to identify novel fusion transcripts in TNBC.MethodsWe analyzed the RNA sequencing data of 360 TNBC samples to identify and filter fusion candidates through SOAPfuse and ChimeraScan analysis. The characteristics, including recurrence, fusion type, chromosomal localization, TNBC subgroup distribution, and clinicopathological correlations, were analyzed in all candidates. Furthermore, we selected the promising fusion transcript and predicted its fusion type and protein coding capacity.ResultsUsing the RNA sequencing data, we identified 189 fusion transcripts in TNBC, among which 22 were recurrent fusions. Compared to para-tumor tissues, TNBC tumor tissues accumulated more fusion events, especially in high-grade tumors. Interestingly, these events were enriched at specific chromosomal loci, and the distribution pattern varied in different TNBC subtypes. The vast majority of fusion partners were discovered on chromosomes 1p, 11q, 19p, and 19q. Besides, fusion events mainly clustered on chromosome 11 in the immunomodulatory subtype and chromosome 19 in the luminal androgen receptor subtype of TNBC. Considering the tumor specificity and frameshift mutation, we selected MFGE8-HAPLN3 as a novel biomarker and further validated it in TNBC samples using PCR and Sanger sequencing. Further, we successfully identified three types of MFGE8-HAPLN3 (E6-E2, E5-E3, and E6-E3) and predicted the ORF of E6-E2, which could encode a protein of 712 amino acids, suggesting its critical role in TNBC.ConclusionsImproved bioinformatic stratification and comprehensive analysis identified the fusion transcript MFGE8-HAPLN3 as a novel biomarker with promising clinical application in the future.

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A fusion of CD63–BCAR4 identified in lung adenocarcinoma promotes tumorigenicity and metastasis

British Journal of Cancer ◽

10.1038/s41416-020-01146-3 ◽

2020 ◽

Vol 124 (1) ◽

pp. 290-298

Author(s):

Kieun Bae ◽

Jin Hee Kim ◽

Hyojik Jung ◽

Sun-Young Kong ◽

Yun-Hee Kim ◽

...

Keyword(s):

Cell Migration ◽

Lung Adenocarcinoma ◽

Fusion Gene ◽

Animal Experiments ◽

Xenograft Model ◽

Fusion Transcript ◽

Sequencing Data ◽

Mouse Xenograft Model

Abstract Background Recently, fusion variants of the breast cancer anti-oestrogen-resistance 4 (BCAR4) gene were recurrently discovered in lung adenocarcinoma from the genome-wide studies. However, the functional characterisation of BCAR4 fusion has not been investigated. Methods Based on the analysis of RNA-sequencing data, we identified a fusion transcript of CD63–BCAR4 in a Korean patient with lung adenocarcinoma who did not harbour any known activating mutations in EGFR and KRAS genes. To investigate the oncogenic effect of CD63–BCAR4, in vitro and in vivo animal experiments were performed. Results In vitro experiments showed strongly enhanced cell migration and proliferation by the exogenous expression of CD63–BCAR4 protein in bronchial epithelial cells. Cell migration was notably reduced after knockdown of BCAR4 fusion by small-interfering RNA. The tumorigenic and metastatic capability of the CD63–BCAR4 fusion was confirmed by using the mouse xenograft model. Fusion-overexpressed cells result in metastasis to the liver and lung as well as the primary tumours after subcutaneous injection into mice. Cyclin D1, MMP1, Slug and mesenchymal markers were significantly increased after CD63–BCAR4 overexpression in the in vitro and in vivo experiments. Conclusions Taken together, our results suggest a newly identified fusion gene, CD63–BCAR4 as a potential novel oncogene in lung adenocarcinoma.

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FusionCatcher - a tool for finding somatic fusion genes in paired-end RNA-sequencing data

10.1101/011650 ◽

2014 ◽

Cited By ~ 118

Author(s):

Daniel Nicorici ◽

Mihaela Satalan ◽

Henrik Edgren ◽

Sara Kangaspeska ◽

Astrid Murumagi ◽

...

Keyword(s):

Real Time ◽

Tumor Cells ◽

Rna Sequencing ◽

Real Time Pcr ◽

Software Tool ◽

Fusion Genes ◽

Sequencing Data ◽

Detection Rates ◽

Somatic Fusion

FusionCatcher is a software tool for finding somatic fusion genes in paired-end RNA-sequencing data from human or other vertebrates. FusionCatcher achieves competitive detection rates and real-time PCR validation rates in RNA-sequencing data from tumor cells. FusionCatcher is available at http://code.google.com/p/fusioncatcher

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Co-fuse: a new class discovery analysis tool to identify and prioritize recurrent fusion genes from RNA-sequencing data

Molecular Genetics and Genomics ◽

10.1007/s00438-018-1454-1 ◽

2018 ◽

Vol 293 (5) ◽

pp. 1217-1229

Author(s):

Sakrapee Paisitkriangkrai ◽

Kelly Quek ◽

Eva Nievergall ◽

Anissa Jabbour ◽

Andrew Zannettino ◽

...

Keyword(s):

Rna Sequencing ◽

Analysis Tool ◽

Fusion Genes ◽

Sequencing Data ◽

New Class ◽

Class Discovery

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Identification of novel mutations of ovarian cancer-related genes from RNA-sequencing data for Japanese epithelial ovarian cancer patients

Endocrine Journal ◽

10.1507/endocrj.ej19-0283 ◽

2020 ◽

Vol 67 (2) ◽

pp. 219-229

Author(s):

Saya Nagasawa ◽

Kazuhiro Ikeda ◽

Kuniko Horie-Inoue ◽

Sho Sato ◽

Satoru Takeda ◽

...

Keyword(s):

Ovarian Cancer ◽

Epithelial Ovarian Cancer ◽

Cancer Patients ◽

Rna Sequencing ◽

Sequencing Data ◽

Novel Mutations

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NUTM1 Is a Rare but Recurrent Fusion Gene Partner in B Cell Precursor Acute Lymphoblastic Leukemia Associated with Increased Expression of Genes on Cytoband 10p12.31

Blood ◽

10.1182/blood-2018-99-112480 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 2823-2823

Author(s):

Femke M. Hormann ◽

Alex Q. Hoogkamer ◽

H. Berna Beverloo ◽

Aurélie Boeree ◽

Ronald W. Stam ◽

...

Keyword(s):

Gene Expression ◽

Acute Lymphoblastic Leukemia ◽

Rna Sequencing ◽

Histone Acetyltransferase ◽

Lymphoblastic Leukemia ◽

Fusion Transcript ◽

Population Based ◽

Fusion Genes ◽

Cell Precursor ◽

Cytogenetic Aberration

Abstract INTRODUCTION In 20-25% of the pediatric B cell precursor acute lymphoblastic leukemia (BCP-ALL) patients, the driving cytogenetic aberration is unknown. It is important to identify more primary lesions in this remaining B-other group to provide better risk stratification and identify possible treatment options. In this study, we aimed to identify novel recurrent fusion genes in BCP-ALL through RNA sequencing. METHODS We used paired-end total RNA Illumina sequencing to detect fusion genes with STAR-fusion and FusionCatcher in a population-based ALL cohort (n=71). We used Affymetrix U133 Plus2 expression arrays in a larger population-based ALL cohort (n=661) and an infant ALL cohort (n=70) to compare gene expression levels. Fluorescent in situ hybridization (FISH) was performed using Cytocell NUTM1 break-apart probe set MPH4800. RESULTS We identified an in-frame SLC12A6-NUTM1 fusion transcript composed of exons 1-2 of SLC12A6 fused to exons 3 to 8 of NUTM1 by RNA sequencing. Both genes are located on 15q14 within 5.3 Kb distance on opposite strands, and the fusion could result from an inversion. The fusion transcript is predicted to encode almost the total NUTM1 protein including the acidic binding domain for the histone acetyltransferase EP300. The SLC12A6-NUTM1 fusion case showed high NUTM1 expression, while NUTM1 expression was absent in the remaining cases. Using gene expression profiling, we identified four additional pediatric and two non-KMT2A-rearranged infant BCP-ALL cases with high NUTM1 expression. In the population-based cohort reflecting all different cytogenetic subtypes, these cases were restricted to the B-other group without known sentinel cytogenetic abnormalities. FISH showed a NUTM1 break apart pattern in all four tested NUTM1-positive cases indicative of a balanced translocation. RNA sequencing confirmed an ACIN1-NUTM1 fusion in one of the infant cases. We conclude that NUTM1 is normally not expressed in leukemic lymphoblasts, and that its expression can be induced by a gene fusion. The karyotypes of the predicted NUTM1 fusion cases combined with RNA sequencing data suggest that different chromosomal rearrangements are involved, likely resulting in different NUTM1 fusion partners. In literature, BRD9-NUTM1, IKZF1-NUTM1, and CUX1-NUTM1 fusions were reported in pediatric B-other cases, and BRD9-NUTM1 and ACIN1-NUTM1 fusions were reported in non-KMT2A-rearranged infants. Our combined aberrant gene expression and FISH results indicate that NUTM1 fusions occur in 2.4% (5/210) of pediatric and in 28% (2/7) of infant BCP-ALL cases without a sentinel cytogenetic aberration. The recurrence of NUTM1 aberrations in BCP-ALL cases without a known driver and the resulting expression of NUTM1 suggests that this fusion could be a new oncogenic driver in leukemia. All seven patients with a NUTM1 fusion achieved continuous complete remission with a median follow-up time of 8.3 years (range 4.8-13.8 years), suggesting that NUTM1 fusions in BCP-ALL have a favorable prognosis. To get an insight in the underlying biology, we compared gene expression between NUTM1-positive and NUTM1-negative pediatric B-other cases. We identified 130 differentially expressed probe sets (FDR ≤0.01) with a peculiar enrichment of those located on chromosome band 10p12.31 (Bonferroni adjusted p=4.05E-04). The genes in cytoband 10p12.31, including BMI1, were variably upregulated in 6/7 NUTM1-positive cases and positively correlated to NUTM1 expression levels. The NUTM1 protein is capable of binding and hereby stimulating the histone acetyltransferase activity of the EP300 protein. The EP300 protein preferentially binds a risk allele of BMI1 associated with increased risk for BCP-ALL. The BMI1 protein has been shown to convert BCR-ABL1-positive progenitor cells into BCR-ABL1-positive BCP-ALL cells. Hence, we postulate that NUTM1 fusion proteins contribute to leukemogenesis by stimulating EP300, leading to upregulation of BMI1 and other 10p12.31 genes in BCP-ALL. CONCLUSION NUTM1 fusions are a rare but recurrent event in BCP-ALL that seems to have a good prognosis. The NUTM1 fusions result in expression of the normally silent NUTM1 gene and are associated with upregulation of a cluster of genes on 10p12.31 including the leukemogenic BMI1 gene. Disclosures No relevant conflicts of interest to declare.

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Partner-independent fusion gene detection by multiplexed CRISPR/Cas9 enrichment and long-read Nanopore sequencing

10.1101/807545 ◽

2019 ◽

Cited By ~ 1

Author(s):

Christina Stangl ◽

Sam de Blank ◽

Ivo Renkens ◽

Tamara Verbeek ◽

Jose Espejo Valle-Inclan ◽

...

Keyword(s):

Fusion Gene ◽

Diagnostic Methods ◽

Partner Choice ◽

Fusion Partner ◽

Nanopore Sequencing ◽

Fusion Genes ◽

Sequencing Data ◽

Gene Detection ◽

Detection Assay ◽

Long Read

AbstractFusion genes are hallmarks of various cancer types and important determinants for diagnosis, prognosis and treatment possibilities. The promiscuity of fusion genes with respect to partner choice and exact breakpoint-positions restricts their detection in the diagnostic setting, even for known and recurrent fusion gene configurations. To accurately identify these gene fusions in an unbiased manner, we developed FUDGE: a FUsion gene Detection assay from Gene Enrichment. FUDGE couples target-selected and strand-specific CRISPR/Cas9 activity for enrichment and detection of fusion gene drivers (e.g. BRAF, EWSR1, KMT2A/MLL) - without prior knowledge of fusion partner or breakpoint-location - to long-read Nanopore sequencing. FUDGE encompasses a dedicated bioinformatics approach (NanoFG) to detect fusion genes from Nanopore sequencing data. Our strategy is flexible with respect to target choice and enables multiplexed enrichment for simultaneous analysis of several genes in multiple samples in a single sequencing run. We observe on average a 508 fold on-target enrichment and identify fusion breakpoints at nucleotide resolution - all within two days. We demonstrate that FUDGE effectively identifies fusion genes in cancer cell lines, tumor samples and on whole genome amplified DNA irrespective of partner gene or breakpoint-position in 100% of cases. Furthermore, we show that FUDGE is superior to routine diagnostic methods for fusion gene detection. In summary, we have developed a rapid and versatile fusion gene detection assay, providing an unparalleled opportunity for pan-cancer detection of fusion genes in routine diagnostics.

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Systematic Identification of Characteristic Genes of Ovarian Clear Cell Carcinoma Compared with High-Grade Serous Carcinoma Based on RNA-Sequencing

International Journal of Molecular Sciences ◽

10.3390/ijms20184330 ◽

2019 ◽

Vol 20 (18) ◽

pp. 4330 ◽

Cited By ~ 5

Author(s):

Saya Nagasawa ◽

Kazuhiro Ikeda ◽

Kuniko Horie-Inoue ◽

Sho Sato ◽

Atsuo Itakura ◽

...

Keyword(s):

Ovarian Cancer ◽

Cell Carcinoma ◽

Rna Sequencing ◽

Functional Annotation ◽

Molecular Mechanisms ◽

Clear Cell ◽

Sequence Data ◽

Clear Cell Carcinoma ◽

Serous Carcinoma ◽

High Grade

Objective: Ovarian cancer has the highest mortality among gynecological cancers. High-grade serous carcinoma (HGSC) is the most common histotype of ovarian cancer regardless of ethnicity, whereas clear cell carcinoma (CCC) is more common in East Asians than Caucasians. The elucidation of predominant signaling pathways in these cancers is the first step towards understanding their molecular mechanisms and developing their clinical management. Methods: RNA sequencing was performed for 27 clinical ovarian specimens from Japanese women. Principal component analysis (PCA) was conducted on the sequence data mapped on RefSeq with normalized read counts, and functional annotation analysis was performed on genes with substantial weights in PCA. Knockdown experiments were conducted on the selected genes on the basis of PCA. Results: Functional annotation analysis of PCA-defined genes showed predominant pathways, such as cell growth regulators and blood coagulators in CCC and transcription regulators in HGSC. Knockdown experiments showed that the inhibition of the calcium-dependent protein copine 8 (CPNE8) and the transcription factor basic helix-loop-helix family member e 41 (BHLHE41) repressed the proliferation of CCC- and HGSC-derived cells, respectively. Conclusions: This study identified CPNE8 and BHLHE41 as characteristic genes for CCC and HGSC, respectively. The systemic identification of differentially expressed genes in CCC and HGSC will provide useful information to understand transcriptomic differences in these ovarian cancers and to further develop potential diagnostic and therapeutic options for advanced disease.

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Suppression of MLL-AF4 by Small Interfering RNAs Inhibits Proliferation and Induces Apoptosis in T(4;11)-Positive SEM Cells.

Blood ◽

10.1182/blood.v104.11.4433.4433 ◽

2004 ◽

Vol 104 (11) ◽

pp. 4433-4433

Author(s):

Johann Greil ◽

Maria Thomas ◽

Christian Huenefeld ◽

Hans-Peter Vornlocher ◽

Philipp Hadwiger ◽

...

Keyword(s):

Fusion Gene ◽

Chromosomal Translocation ◽

Fusion Transcript ◽

Derivative Chromosome ◽

Mrna Levels ◽

Small Interfering Rnas ◽

Chromosome 11 ◽

Inhibition Of Proliferation ◽

Treatment Concepts ◽

Cell Cycle Transition

Abstract The reciprocal chromosomal translocation t(4;11)(q21;q23) creates the fusion genes MLL-AF4 and AF4-MLL located on derivative chromosome 11 or derivative chromosome 4, respectively. We used small interfering RNAs to suppress the MLL-AF4 fusion gene in the t(4;11)-positive leukaemic cell line SEM. Electroporation of SEM cells with MLL-AF4 siRNAs caused a more than two fold transient decrease in MLL-AF4 mRNA levels, which lasted for three days. The reduction in MLL-AF4 fusion transcript levels were associated with a severely diminished clonogenicity, inhibition of proliferation and of G1-S cell cycle transition and induction of apoptosis. Therefore, MLL-AF4 siRNAs are not only useful to study the functions of MLL-AF4 in leukaemogenesis, but may be also promising agents for novel treatment concepts for t(4;11)-associated leukaemias.

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