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Ferrata Storti Foundation
Haematologica 2022 Volume 107(1):100-111
Fusion gene detection by RNA-sequencing complements diagnostics of acute myeloid leukemia and identifies recurring NRIP1-MIR99AHG rearrangements
Paul Kerbs,1,2,3 Sebastian Vosberg,1,2,3 Stefan Krebs,4 Alexander Graf,4 Helmut Blum,4 Anja Swoboda,1 Aarif M. N. Batcha,5 Ulrich Mansmann,5 Dirk Metzler,6 Caroline A. Heckman,7 Tobias Herold1,2,3 and Philipp A. Greif1,2,3
1Department of Medicine III, University Hospital, LMU Munich, Munich, Germany; 2German Cancer Consortium (DKTK), partner site Munich, Germany; 3German Cancer Research Center (DKFZ), Heidelberg, Germany; 4Laboratory for Functional Genome Analysis (LAFUGA), Gene Center, LMU Munich, Munich, Germany; 5Department of Medical Data Processing, Biometry and Epidemiology, LMU Munich, Munich, Germany; 6Division of Evolutionary Biology, Faculty of Biology, LMU Munich, Planegg-Martinsried, Germany and 7Institute for Molecular Medicine Finland (FIMM), University of Helsinki, Helsinki, Finland
ABSTRACT
Identification of fusion genes in clinical routine is mostly based on cyto- genetics and targeted molecular genetics, such as metaphase karyotyp- ing, fluorescence in situ hybridization and reverse-transcriptase poly- merase chain reaction. However, sequencing technologies are becoming more important in clinical routine as processing time and costs per sample decrease. To evaluate the performance of fusion gene detection by RNA- sequencing compared to standard diagnostic techniques, we analyzed 806 RNA-sequencing samples from patients with acute myeloid leukemia using two state-of-the-art software tools, namely Arriba and FusionCatcher. RNA-sequencing detected 90% of fusion events that were reported by rou- tine with high evidence, while samples in which RNA-sequencing failed to detect fusion genes had overall lower and inhomogeneous sequence cover- age. Based on properties of known and unknown fusion events, we devel- oped a workflow with integrated filtering strategies for the identification of robust fusion gene candidates by RNA-sequencing. Thereby, we detected known recurrent fusion events in 26 cases that were not reported by rou- tine and found discrepancies in evidence for known fusion events between routine and RNA-sequencing in three cases. Moreover, we identified 157 fusion genes as novel robust candidates and comparison to entries from ChimerDB or Mitelman Database showed novel recurrence of fusion genes in 14 cases. Finally, we detected the novel recurrent fusion gene NRIP1- MIR99AHG resulting from inv(21)(q11.2;q21.1) in nine patients (1.1%) and LTN1-MX1 resulting from inv(21)(q21.3;q22.3) in two patients (0.25%). We demonstrated that NRIP1-MIR99AHG results in overexpression of the 3' region of MIR99AHG and the disruption of the tricistronic miRNA cluster miR-99a/let-7c/miR-125b-2. Interestingly, upregulation of MIR99AHG and deregulation of the miRNA cluster, residing in the MIR99AHG locus, are known mechanisms of leukemogenesis in acute megakaryoblastic leukemia. Our findings demonstrate that RNA-sequencing has a strong potential to improve the systematic detection of fusion genes in clinical applications and provides a valuable tool for fusion discovery.
Introduction
Fusion genes result from chromosomal aberrations, such as translocations, dupli- cations, inversions or small interstitial deletions. On the transcript level, fusion genes may not only reflect underlying genomic rearrangements but may also arise due to
Acute Myeloid Leukemia
Presented in part at the 61st ASH Annual Meeting Kerbs P, et al. Blood. 2019;134(Suppl_1):4655.
Correspondence:
PHILIPP A. GREIF
pgreif@med.lmu.de
Received: January 26, 2021. Accepted: May 3, 2021. Pre-published: June 17, 2021.
https://doi.org/10.3324/haematol.2021.278436 ©2022 Ferrata Storti Foundation
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