Page 223 - Haematologica April 2020

MMEJ drives 8q24 rearrangements in myeloma
cations in MM, and the IGH-MYC primary events in Burkitt’s lymphoma, are mediated by AID and class switch recombination.2,4,41 Therefore, the IGH-MYC rearrangements may occur through an as yet unknown AID-independent mechanism.
The mechanism driving MYC rearrangements is likely not to involve NHEJ, which would result in blunt ended rearrangements.33 We have shown that MYC rearrange- ments are more likely to have short homologous sequences in common to both partner chromosomes, which is not seen as frequently in the primary IGH translocations. Short homologous sequences are indicative of MMEJ,42 rather than NHEJ, and result from fork stalling and template switching during DNA replication or through microhomology-mediated break induced repair.43,44 The proteins involved in MMEJ include PARP1, Rad50, and Ercc1, whereas MMEJ is inhibited by function- al ATM, H2AX, 53BP1, and BRCA1.42 We have previously shown that mutation of ATM, BRCA1 and other genes involved in DNA homologous recombination are associat- ed with increased levels of loss of heterozygosity in MM patients.45 Itislikelythatdisruptionofthispathwayiskey to genomic instability and progression of disease.
The non-Ig chromosomal partners of MYC are not ran- dom and are known to contain super-enhancer elements.5,6 From our analysis of the breakpoints at the most frequent non-Ig locations [6p24.3 (TXNDC5/BMP6), 1p12 (FAM46C), 6q21 (FOXO3)], we show that the breakpoints at these genes are also clustered. The breakpoints are, in general, contained within TAD which are more likely to interact with one another.46,47 Each TAD at the partner chromosome contains a super-enhancer and breakpoints rarely fall outside of the TAD. The rearrangements are predicted to result in a changed TAD structure that places MYC in the same domain as the super-enhancer from the partner locus. If breakpoints were to occur outside of the TAD with the super-enhancer, there would be a lower likelihood of it interacting with MYC and expression would not be enhanced.
We identified 149 partner loci for MYC rearrangements, but 67.2% of the samples with translocations involve one of the six main partners. The Ig partners have strong super-enhancers in MM, but there are many other active super-enhancers and so it is likely that these six main part- ners are constrained by chromatin structure. The break- points at 8q24 surround an epigenetically active region, defined by the active chromatin marks H3K27Ac, H3K36me3 and H3K4me1, as well as DNaseI hypersensi- tivity sites. It may be that epigenetically active, and there-
fore accessible, loci are preferred translocation partners,48,49 and the nuclear localization of chromosomes may play a part, too.50
Each of these different rearrangements results in overex- pression of MYC. MYC is not the only gene at 8q24, and, indeed, PVT1 is significantly over-expressed in our dataset. PVT1 is a long non-coding RNA and is associated with inhibition of apoptosis and increased proliferation.51 It has also been shown that PVT1 interacts with MYC, resulting in a stable protein, and that ablation of PVT1 results in diminished tumorigenicity.52 It may be that the gene complex encompassing MYC and PVT1 is required for oncogenesis and merits further study.
Besides PVT1, we also identified other genes that are direct targets of c-Myc and are over-expressed in 8q24- rearranged samples. These included HK2, a key enzyme involved in glucose metabolism. It has previously been shown that silencing of HK2 sensitizes cancer cells to other drugs, and so overexpression of HK2 in MYC-rearranged myeloma may be a key drug resistance mechanism.53 Additional genes involved in important cellu- lar functions that increase the oncogenic potential of myelo- ma cells were also identified, such as ribosome biosynthesis and translation initiation; these are likely to contribute to the poor prognosis seen in MYC-rearranged myeloma.5,14 Targeting MYC could, therefore, be an effective way to dis- rupt many essential tumor features in one hit.
This study provides evidence of complex chromosomal rearrangements at 8q24 as a key cause of MYC oncogenic upregulation. Although we found that several MYC abnormalities are associated with prognosis in this dataset, including MYC-IGL and complex translocations, we have previously shown that the association is not independent of other genomic and clinical markers.54 However, it may be possible that, with longer follow up, MYC abnormalities may be independently associated with overall survival and be a marker of poor outcome. We also show a specific pattern of chromosomal break- points suggesting the role of the chromatin landscape in tumorigenesis. The mechanism of DNA breaks clearly differs between MYC rearrangements, resulting from MMEJ rather than NHEJ, and differs in myeloma com- pared to primary MYC translocations in lymphoma.
Funding
Funding support for the CoMMpass dataset was provided by the Myeloma Genome Project. The CoMMpass dataset was generated by the Multiple Myeloma Research Foundation in col- laboration with the Multiple Myeloma Research Consortium.
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