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. 2017 Jan;102(1):118-129.
doi: 10.3324/haematol.2016.151035. Epub 2016 Sep 15.

ZNF384-related Fusion Genes Define a Subgroup of Childhood B-cell Precursor Acute Lymphoblastic Leukemia With a Characteristic Immunotype

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ZNF384-related Fusion Genes Define a Subgroup of Childhood B-cell Precursor Acute Lymphoblastic Leukemia With a Characteristic Immunotype

Shinsuke Hirabayashi et al. Haematologica. .
Free PMC article

Abstract

Fusion genes involving ZNF384 have recently been identified in B-cell precursor acute lymphoblastic leukemia, and 7 fusion partners have been reported. We further characterized this type of fusion gene by whole transcriptome sequencing and/or polymerase chain reaction. In addition to previously reported genes, we identified BMP2K as a novel fusion partner for ZNF384 Including the EP300-ZNF384 that we reported recently, the total frequency of ZNF384-related fusion genes was 4.1% in 291 B-cell precursor acute lymphoblastic leukemia patients enrolled in a single clinical trial, and TCF3-ZNF384 was the most recurrent, with a frequency of 2.4%. The characteristic immunophenotype of weak CD10 and aberrant CD13 and/or CD33 expression was revealed to be a common feature of the leukemic cells harboring ZNF384-related fusion genes. The signature gene expression profile in TCF3-ZNF384-positive patients was enriched in hematopoietic stem cell features and related to that of EP300-ZNF384-positive patients, but was significantly distinct from that of TCF3-PBX1-positive and ZNF384-fusion-negative patients. However, clinical features of TCF3-ZNF384-positive patients are markedly different from those of EP300-ZNF384-positive patients, exhibiting higher cell counts and a younger age at presentation. TCF3-ZNF384-positive patients revealed a significantly poorer steroid response and a higher frequency of relapse, and the additional activating mutations in RAS signaling pathway genes were detected by whole exome analysis in some of the cases. Our observations indicate that ZNF384-related fusion genes consist of a distinct subgroup of B-cell precursor acute lymphoblastic leukemia with a characteristic immunophenotype, while the clinical features depend on the functional properties of individual fusion partners.

Figures

Figure 1.
Figure 1.
Flow chart of the analysis of patients. The respective numbers of patients and cohorts that were investigated are presented in a hierarchical fashion. See also the Online Supplementary Information and Online Supplementary Table S1. ALL: acute lymphoblastic leukemia; BCP-ALL: B-cell precursor ALL; B-others: BCP-ALL without conventional genetic abnormalities; RNA: ribonucleic acid; RNA-seq-Ex1 and Ex2: RNA-sequencing experiment 1 and 2; RT-PCR-Ex1 and Ex2: reverse transcription polymerase chain reaction experiment 1 and 2; Ex: experiment.
Figure 2.
Figure 2.
Structure of the ZNF384-related fusions. Structures of fusion proteins and nucleotide sequences of fusion points of (A) TCF3-ZNF384, (B) TAF15-ZNF384, (C) CREBBP-ZNF384, and (D) BMP2K-ZNF384 are presented. Exon numbers and boundaries are marked below the protein structures. The black arrowhead shows the donor site fusion point. The white arrowhead shows the acceptor site fusion point. Five and four different in-frame fusions for TCF3-ZNF384 and CREBBP-ZNF384, respectively, are depicted with the nucleotide sequence, predicted amino acids, and chromatogram. The patients in whom a particular fusion isoform was found are indicated on the right. Ex: exon; TAZ2: transcription adaptor putative zinc finger; KIX: kinase-inducible domain interacting; RING: really interesting new gene; PHD: plant homeodomain; CREB: cyclic adenosine monophosphate (AMP) response element binding protein; RNA: ribonucleic acid; ZN: zinc finger; RAN: Ras-related nuclear proteins; BMP-2: bone morphogenetic protein 2.
Figure 3.
Figure 3.
Immunophenotypic characteristics of B-cell precursor acute lymphoblastic leukemia (BCP-ALL) patients with ZNF384-related fusion genes. (A) Typical histograms of CD19, CD10, aberrant myeloid antigens (CD13 and CD33), and CD66c of ZNF384-related fusion gene-positive patients and TCF3-PBX1 patients are indicated with a positive rate (%). X-axis, fluorescence intensity; Y-axis, relative cell number. (B) The positivity (percentage) of CD10, 13, and 33 of ZNF384-related fusion gene-positive BCP-ALL patients (22 cases, excepting EP300-ZNF384) and TCF3-PBX1-positive or wild-type ZNF384 BCP-ALL patients was plotted on a scattergram. The detailed list of positivity for each immunophenotypic marker of the patients is presented in the Online Supplementary Table S5. ZNF384-WT indicates cases without ZNF384 rearrangement.
Figure 4.
Figure 4.
Characteristics of gene expression profile in TCF3-ZNF384-positive acute lymphoblastic leukemia (ALL). (A) Two-way hierarchical clustering was performed on filtered microarray probes, those up- or downregulated by 2-fold or more in TCF3-ZNF384- (red, n=10), in comparison with TCF3-PBX1-positive (blue, n=19) ALL. The results are displayed using a heat map as a dendrogram. (B) Among the genes that exhibited a 2 times higher or lower expression in either TCF3-ZNF384- and TCF3-PBX1-positive ALL (listed in the Online Supplementary Table S6), the expressions of PAX5 and VPREB genes measured by microarray analysis were plotted on a scatter diagram, and bars representing the mean±SD are presented. The data of wild-type ZNF384 B-cell precursor acute lymphoblastic leukemia (BCP-ALL) patients are also presented. (C) Gene set enrichment analysis (GSEA) for curated gene sets of hematopoietic precursors was performed on the differentially expressed genes between TCF3-ZNF384- (red) and TCF3-PBX1-positive (blue) ALL. Enrichment plots for the hematopoietic stem cells (HSCs), multi-lymphoid progenitor (MLP), pro-B cell (Pro-B), early T-cell precursors (ETP), common myeloid progenitors (CMP), granulocyte-monocyte progenitor (GMP), and megakaryocyte-erythroid progenitor cell (MEP) signatures are presented. Bold lines represent significant enrichments {false discovery rate (FDR) q-value<0.25 and/or nominal (NOM) P-value<0.05)}. NES: normalized enrichment score.
Figure 5.
Figure 5.
Outcomes of patients with ZNF384-related fusion genes. (A) Kaplan-Meier estimates of event-free survival (EFS) for patients with TCF3-ZNF384, EP300-ZNF384, and wild-type ZNF384 (log-rank P=0.35 in wild-type ZNF384 vs. TCF3-ZNF384, log-rank P=0.17 in TCF3-ZNF384 vs. EP300-ZNF384. (B) Overall survival (OS) for the same as above (log-rank P=0.15 in wild-type ZNF384 vs. TCF3-ZNF384, log-rank P=0.14 in TCF3-ZNF384 vs. EP300-ZNF384.

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