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RNA-seq identifies clinically relevant fusion genes in leukemia including a novel MEF2D/CSF1R fusion responsive to imatinib

Leukemia volume 28, pages 977–979 (2014)Cite this article

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To date, more than 500 fusion genes have been described in hematological malignancies, yet only a small fraction of these are sufficiently common, and with a known clinical impact, to justify routine clinical analysis.1 For this small subgroup, however, their presence can provide crucial information for diagnosis, prognosis and treatment. For example, the presence of fusion genes such as BCR/ABL1 in chronic myeloid leukemia (CML) and PML/RARA in acute promyelocytic leukemia warrants treatment specifically targeting the fusion proteins.2, 3 Other gene fusions, for which there might not exist targeted therapies, are used to risk stratify leukemias into different treatment groups; for example, ETV6/RUNX1, TCF3/PBX1 and various MLL fusions in childhood B-cell acute lymphoblastic leukemia (ALL).4 The proportion of cases harboring known clinically relevant fusion genes varies substantially between hematological malignancies, but in ALL and acute myeloid leukemia (AML) 25–45% of the cases are positive for any of the clinically relevant fusion genes.4, 5 To what extent the remaining cases harbor novel fusions, perhaps also suitable for targeted treatment, is not known, largely because the process of identifying new gene fusions has been laborious.

Here, we describe how RNA sequencing (RNA-seq) can be used as a reliable tool to identify known clinically relevant fusion genes, in addition to novel fusions. Fifteen leukemic cell lines that were derived from ALL, AML or CML containing a known prognostically relevant fusion gene were included in the study (Table 1). In addition to these, one primary preB-ALL, negative for BCR/ABL1, TCF3/PBX1 and ETV6/RUNX1, but harboring a t(1;5)(q21;q33), was studied (for a detailed case history, see Supplementary Materials and Methods). Massively parallel sequencing was performed on mRNA libraries from all samples, generating between 12 and 21 million read pairs for each library (Supplementary Table S1 and Supplementary Materials and Methods). A list of 42 candidate fusion genes was identified using chimerascan7 followed by a filtering approach. Of these, 31 (74%) could be verified as genuine gene fusions using reverse transcriptase–PCR and Sanger sequencing (Table 1). Notably, this included all 15 clinically relevant fusion genes previously known to be present in the cell lines. Seven of the remaining verified fusions were reciprocal to previously known fusions and nine were novel. Of the nine novel fusions, seven were expected to produce nonfunctional proteins or no protein, either because the translated regions were fused out of frame or because of fusion to untranslated regions. However, these nonfunctional fusions contained genes such as RUNX1, NUP214 and EBF (Table 1) that are known to be involved in leukemogenesis in several types of aberrations, for example, in-frame fusions.1 Hence, it seems as if the establishment of truncating fusion genes is an alternative way to impair the function of important genes during leukemogenesis. Similar out-of-frame fusion genes were recently shown to be present in 31 of 200 AMLs (16%).8

Table 1 Fusion genes detected by RNA sequencing
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Acknowledgements

This work was supported by the Swedish Cancer Society, the Swedish Children’s Cancer Foundation, the Inga-Britt and Arne Lundberg Foundation, the Gunnar Nilsson Cancer Foundation, the Medical Faculty of Lund University and the Swedish Research Council (personal project grant to TF, Hemato-Linné and BioCARE strategic research program grants).

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  1. Department of Clinical Genetics, University and Regional Laboratories, Lund University, Lund, Sweden

    H Lilljebjörn, H Ågerstam, C Orsmark-Pietras, M Rissler, H Ehrencrona & T Fioretos

  2. Department of Hematology and Coagulation, Skåne University Hospital, Lund, Sweden

    L Nilsson & J Richter

  3. Department of Molecular Medicine and Gene Therapy, Lund University, Lund, Sweden

    J Richter

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Lilljebjörn, H., Ågerstam, H., Orsmark-Pietras, C. et al. RNA-seq identifies clinically relevant fusion genes in leukemia including a novel MEF2D/CSF1R fusion responsive to imatinib. Leukemia 28, 977–979 (2014). https://doi.org/10.1038/leu.2013.324

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