SVAtools for junction detection of genome-wide chromosomal rearrangements by mate-pair sequencing (MPseq)

Johnson, Sarah H.; Smadbeck, James B.; Smoley, Stephanie A.; Gaitatzes, Athanasios; Murphy, Stephen J.; Harris, Frances; Drucker, Travis; Zenka, Roman M.; Pitel, Beth A.; Rowsey, Ross; Hoppman, Nicole L.; Aypar, Umut; Sukov, William R.; Jenkins, Robert B.; Feldman, Andrew L.; Kearney, Hutton M.; Vasmatzis, George

doi:10.1016/j.cancergen.2017.11.009

Cited by 65 publications

(82 citation statements)

References 26 publications

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“…To date, the full characterization of KMT2A/MLLT10 gene fusions has heavily relied upon a combination of conventional chromosome, FISH, and reverse transcriptase‐polymerase chain reaction studies . To further characterize 2 cryptic KMT2A/MLLT10 fusions with “normal” chromosomes studies (patients 18 and 48), we performed MPseq, a next‐generation sequencing (NGS)‐based assay that resolves chromosomal rearrangements with significantly higher resolution and precision compared to traditional cytogenetic methodoliges . MPseq results for patient 18 detected an insertion of exons 9‐24 of MLLT10 into intron 9 of KMT2A (illustrated in Aypar et al) and predicts an in frame chimeric KMT2A/MLLT10 fusion.…”

Section: Discussionmentioning

confidence: 99%

“…This optimization is made possible during library preparation by the generation of 2‐5 kilobase fragments that have 101 base pairs sequenced on each fragment end, resulting in a lower read depth necessary to detect structural variants in the genome. Detailed protocols and additional information regarding MPseq technology and bioinformatics tools can be found in Drucker et al, Johnson et al, and Aypar et al…”

Section: Methodsmentioning

confidence: 99%

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Acute leukemias harboring KMT2A/MLLT10 fusion: a 10‐year experience from a single genomics laboratory

Peterson

Sukov

Pitel

et al. 2019

Genes Chromosomes & Cancer

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The MLLT10 (formerly AF10) gene is the fourth most common KMT2A fusion partner across all acute leukemias and requires at least 3 breaks to form an in‐frame KMT2A/MLLT10 fusion due to the opposite orientation of each gene. A 10‐year retrospective review was performed to identify individuals from all age groups that harbor KMT2A/MLLT10 fusion obtained by our KMT2A/MLLT10 dual‐color dual‐fusion fluorescence in situ hybridization (D‐FISH) assay. Of the 60 unique individuals identified, 31 were male and 29 were female (M:F ratio, 1.1:1) with ages ranging from 3 days to 86 years (mean 21.5 years, median 5.5 years). The diagnoses included acute myeloid leukemia (AML) (49 patients, 82%), B‐ or T‐lymphoblastic leukemia/lymphoma (7 patients, 12%), myeloid sarcoma (3 patients, 5%), and a single case (2%) of undifferentiated leukemia. Twenty‐seven of 49 patients (55%) with AML were in the infant or pediatric age group. Fifty‐three of 60 patients (88%) had KMT2A/MLLT10 D‐FISH signal patterns mostly consisting of single fusions. In addition, 10 (26%) of 38 patients with conventional chromosome studies had “normal” (5 patients) or abnormal (5 patients) chromosome studies that lacked structural or numeric abnormalities involving chromosomes 10 or 11, implying cryptic cytogenetic mechanisms for KMT2A/MLLT10 fusion. Lastly, mate‐pair sequencing was performed on 4 AML cases, 2 of which had “normal” chromosome studies and cryptic KMT2A/MLLT10 fusion as detected by KMT2A/MLLT10 D‐FISH studies, and verified the multiple breaks required to generate KMT2A/MLLT10 fusion.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Acute leukemias harboring KMT2A/MLLT10 fusion: a 10‐year experience from a single genomics laboratory

Peterson

Sukov

Pitel

et al. 2019

Genes Chromosomes & Cancer

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“…Mate pair sequencing (MPseq) was performed as described previously . Briefly, DNA was isolated using the Puregene extraction protocol (Qiagen).…”

Section: Methodsmentioning

confidence: 99%

“…Alignment is performed using the BIMAv3 alignment method . The mapped sequences are then analyzed using SVAtools, which consists of a breakpoint junction detection step and a copy number variant detection step …”

Section: Methodsmentioning

confidence: 99%

RNA sequencing identifies a novel USP9X‐USP6 promoter swap gene fusion in a primary aneurysmal bone cyst

Blackburn

Davila

Jackson

et al. 2019

Genes Chromosomes & Cancer

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Primary aneurysmal bone cyst (ABC) is a benign multiloculated cystic lesion of bone that is defined cytogenetically by USP6 gene rearrangements. Rearrangements involving USP6 are promoter swaps, usually generated by fusion of the noncoding upstream exons of different partner genes with exon 1 or 2 of USP6, thus leading to transcriptional upregulation of full‐length USP6 coding sequence. Testing for USP6 rearrangements is used diagnostically to distinguish it from secondary ABC and other giant cell‐rich primary bone tumors. In this report, we present a case of a 16‐year‐old male with a primary ABC of the left distal femur. USP6 break apart fluorescence in situ hybridization was positive for a rearrangement and conventional chromosome analysis identified a reciprocal X;17 translocation. In order to identify the putative USP6 fusion partner, we performed RNA sequencing and uncovered a novel USP9X‐USP6 promoter swap fusion. This result was confirmed by reverse transcription‐polymerase chain reaction (RT‐PCR) and by mate pair sequencing thus showing the utility of these alternative methodologies in identifying novel fusion candidates. Ubiquitin‐specific protease 9X (USP9X), like USP6, encodes a highly conserved substrate‐specific deubiquitylating enzyme. USP9X is highly expressed in a number of tissue types and acts as both an oncogene and tumor suppressor in several human cancers. We conclude that oncogenic activation of USP6 via USP9X promoter exchange represents a novel driver of primary ABC formation.

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“…Junctions and CNVs were graphically illustrated using genome, junction and region plots as described in Johnson, et al 7 We re- Figure 2). This restriction was necessary for the validation of the MPseq assay by comparing it to the gold standard FISH assay and FISH probes were available for the above-listed targets.…”

Section: Structural Variation Visualizationmentioning

confidence: 99%

Mate pair sequencing improves detection of genomic abnormalities in acute myeloid leukemia

Aypar

Smoley

Pitel

et al. 2018

European J of Haematology

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Objective: Acute myeloid leukemia (AML) can be subtyped based on recurrent cytogenetic and molecular genetic abnormalities with diagnostic and prognostic significance. Although cytogenetic characterization classically involves conventional chromosome and/or fluorescence in situ hybridization (FISH) assays, limitations of these techniques include poor resolution and the inability to precisely identify breakpoints. Method:We evaluated whether an NGS-based methodology that detects structural abnormalities and copy number changes using mate pair sequencing (MPseq) can enhance the diagnostic yield for patients with AML.Results: Using 68 known abnormal and 20 karyotypically normal AML samples, each recurrent primary AML-specific abnormality previously identified in the abnormal samples was confirmed using MPseq. Importantly, in eight cases with abnormalities that could not be resolved by conventional cytogenetic studies, MPseq was utilized to molecularly define eight recurrent AML-fusion events. In addition, MPseq uncovered two cryptic abnormalities that were missed by conventional cytogenetic studies. Thus, MPseq improved the diagnostic yield in the detection of AML-specific structural rearrangements in 10/88 (11%) of cases analyzed. Conclusion:Utilization of MPseq represents a precise, molecular-based technique that can be used as an alternative to conventional cytogenetic studies for newly diagnosed AML patients with the potential to revolutionize the diagnosis of hematologic malignancies. K E Y W O R D Sacute myeloid leukemia, molecular cytogenetics, MPseq

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SVAtools for junction detection of genome-wide chromosomal rearrangements by mate-pair sequencing (MPseq)

Cited by 65 publications

References 26 publications

Acute leukemias harboring KMT2A/MLLT10 fusion: a 10‐year experience from a single genomics laboratory

Acute leukemias harboring KMT2A/MLLT10 fusion: a 10‐year experience from a single genomics laboratory

RNA sequencing identifies a novel USP9X‐USP6 promoter swap gene fusion in a primary aneurysmal bone cyst

Mate pair sequencing improves detection of genomic abnormalities in acute myeloid leukemia

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