Structure and evolution of double minutes in diagnosis and relapse brain tumors

Abstract

Double minute chromosomes are extrachromosomal circular DNA fragments frequently found in brain tumors. To understand their evolution, we characterized the double minutes in paired diagnosis and relapse tumors from a pediatric high-grade glioma and four adult glioblastoma patients. We determined the full structures of the major double minutes using a novel approach combining multiple types of supporting genomic evidence. Among the double minutes identified in the pediatric patient, only one carrying EGFR was maintained at high abundance in both samples, whereas two others were present in only trace amounts at diagnosis but abundant at relapse, and the rest were found either in the relapse sample only or in the diagnosis sample only. For the EGFR-carrying double minutes, we found a secondary somatic deletion in all copies at relapse, after erlotinib treatment. However, the somatic mutation was present at very low frequency at diagnosis, suggesting potential resistance to the EGFR inhibitor. This mutation caused an in-frame RNA transcript to skip exon 16, a novel transcript isoform absent in EST database, as well as about 700 RNA-seq of normal brains that we reviewed. We observed similar patterns involving longitudinal copy number shift of double minutes in another four pairs (diagnosis/relapse) of adult glioblastoma. Overall, in three of five paired tumor samples, we found that although the same oncogenes were amplified at diagnosis and relapse, they were amplified on different double minutes. Our results suggest that double minutes readily evolve, increasing tumor heterogeneity rapidly. Understanding patterns of double minute evolution can shed light on future therapeutic solutions to brain tumors carrying such variants.Electronic supplementary materialThe online version of this article (10.1007/s00401-018-1912-1) contains supplementary material, which is available to authorized users.

Enhancer hijacking determines intra- and extrachromosomal circularMYCNamplicon architecture in neuroblastoma

“…We extend this observation to homogeneously staining regions, which form extremely expanded stretches of chromatin in interphase nuclei and lose chromosomal territoriality 25 Reconstruction of amplicons has previously relied on combining structural breakpoint coordinates to infer the underlying structure. This regularly resulted in ambiguous amplicon reconstructions, which had to be addressed by secondary data such as Chromium linked reads or optical mapping 4,6,24 . We demonstrate the feasibility of long-read de novo assembly for the reconstruction of amplified genomic neighborhoods.…”

“…Over time, amplified DNA acquires additional internal rearrangements as well as coding mutations, which can confer adaptive advantages such as resistance to targeted therapy [5][6][7] .…”

Comprehensive molecular characterization of pediatric treatment-induced high-grade glioma: A distinct entity despite disparate etiologies with defining molecular characteristics and potential therapeutic targets

“…Amplification was confirmed in selected cases by fluorescence in situ hybridization, which demonstrated a double minute pattern, a finding typical of episomal amplification (Fig. 4C) 16 . There was a trend toward an increased frequency of chromothripsis in cases with a coincident pathogenic TP53 mutation (5/8 cases, 62.5%) relative to those with wild-type TP53 (0/4 cases, 0%, p=0.08).…”

ATAC-seq identifies thousands of extrachromosomal circular DNA in cancers and cell lines

“…Many of the abundant eccDNA loci intersect with unprocessed pseudogenes, which are known to have introns and regulatory sequences, but crippled by stop codons in the open reading frames (Tutar, 2012). Since eccDNA evolve and pick up substitution, insertion and deletion mutations (Turner et al, 2017;Xu et al, 2019), it is tempting to speculate that amplification of unprocessed pseudogenes on eccDNA and their evolution may make these genes translationally competent to give an unknown advantage during tumorigenesis.…”

“…This somatic copy number variation is present in 43% of GBM patients (Maire and Ligon, 2014). Recent studies have provided further evidence that this oncogenic amplification occurs on eccDNA (deCarvalho et al, 2018;Turner et al, 2017;Xu et al, 2019). To check if we can detect the eccDNA in ATAC-seq data generated from GBM cell lines we turned to six ATAC-seq libraries generated from GBM cell lines developed from a single glioblastoma patient (Xie et al, 2018).…”