Hi-C analysis of amplification of MYC, PVT1, and CCDC26 on marker chromosomes in the NB-4 cell line.

MYC proto-oncogene may be amplified ectopically in acute myeloid leukemia (AML) as extrachromosomal DNA (ecDNA) such as double minutes (dmins). However, the mechanism and location of MYC amplification have not been fully elucidated. To characterize the location of MYC and its surrounding structure in NB-4 cells, we conducted this study using in situ high-throughput chromosomal conformation capture (in situ Hi-C). Hi-C analysis was performed in NB-4 cell line. Whole genome sequencing (WGS) and fluorescence in situ hybridization (FISH) were used for confirmation. Hi-C revealed an inversion involving PVT1 and CCDC26 and amplified segments involving MYC, PVT1, and CCDC26 on 8q24.21 region. MYC, PVT1, and CCDC26 were found not only on chromosome 8, but also somewhere intranuclear, other than on chromosome 8. Karyotyping revealed only two normal chromosomes 8, and the others were missing or abnormal. FISH revealed the presence of MYC, PVT1, and CCDC26 on the two normal chromosomes 8 and multiple marker chromosomes. Our results suggest that the numerical and structural abnormalities of chromosome 8 precede MYC amplification and moving. MYC may have properties that move not only to dmins, but also to other chromosomes such as marker chromosomes for unknown but certain reasons. MYC ectopic amplification is not only a phenomenon in solid tumors, but also a recurrent phenomenon in AML. Furthermore, in a broader sense, ectopic amplification is a form of abnormal oncogenes. We propose Hi-C as a screening method for this phenomenon. We will further verify this phenomenon in various phases of multiple AML cell lines and patient samples.