Novel Fusion Transcripts PVT1-NSMCE2 and BF104016-NSMCE2 Resulting In Depletion Of NSMCE2 Protein In Acute Myeloid Leukemia Harboring 8q24 Amplicons

Little is known about the molecular pathogenesis of acute myeloid leukemia (AML) with high-level DNA amplifications commonly defined as the presence of double minutes (dmins) and/or homogeneously staining regions (hsr), although dmins and hsr are found in nearly 1% of karyotypically abnormal AML and myelodysplastic syndromes (MDS). It is sometimes difficult to precisely assign dmins, hsr, and marker chromosomes (mars) to specific chromosomal bands, while most of them consist of an amplified segment from chromosome band 8q24 or 11q23. In AML, the 8q24 amplicon has been previously identified in a commonly involved segment of about 4 Mb, which contains several genes such as TRIB1 (C8FW), NSMCE2 (non-SMC element 2), PVT1, and MYC. To further define the pathogenetic lesions of 8q24 amplicons in AML, we performed comprehensive molecular studies on leukemic cells from a patient with AML (patient 1) with two mars and leukemic cell lines HL60 and K562, identifying two novel chimeric transcripts, PVT1-NSMCE2 and BF104016-NSMCE2. Regarding PVT1-NSMCE2 fusion transcripts, PVT1 exon 1a fused to NSMCE2 exon 3 in patient 1, PVT1 exon 3a to NSMCE2 exon 4 in patient 2, and PVT1 exon 4b to NSMCE2 exon 4 in K562. Patient 2 was identified from the screening of an additional 50 patients with AML or MDS by RT-PCR. As for BF104016-NSMCE2 fusion transcript, BF104016 exon 1 fused to NSMCE2 exon 6 in HL60 harboring hsr and dmins. BF104016 is located inside the CCDC26 gene, sharing the same exon of CCDC26. In patient 1, fluorescence in situ hybridization identified the amplification of 5’PVT1 on the mars and dmins, and the amplification of NSMCE2 only on the mars. In HL60, PVT1 and NSMCE2 were amplified on dmins. Although real-time quantitative PCR showed the amplification of the aberrant NSMCE2 chimeric transcripts, western blot analysis demonstrated the depletion of NSMCE2 protein. High-resolution oligonucleotide array analysis demonstrated two amplicons at 8q24 commonly found in patient 1 and HL60. Furthermore, the coding directions of these three fusion genes were different at the 8q24; NSMCE2 and PVT1 are transcribed from centromere toward telomere, opposite to the direction of transcription of the CCDC26 gene. These results suggest that the chimeric formation of these genes were caused by chromothripsis. PVT1 and CCDC26 were known as large intervening non-coding RNAs, both of which were considered to be associated with the oncogenesis. NSMCE2 is known as a small ubiquitin-like modifier (SUMO) E3 ligase and is required for DNA repair. Functional analysis of NSMCE2 chimeric transcripts with PVT1 or CCDC26 will contribute to understanding of the leukemogenesis in AML harboring 8q24 amplicons.