A recent article published by Heliyon on CellPress provides an overview of the source, structure, and function of ecDNA, while compiling recent advancements in ecDNA in the field of cancers. To ensure a comprehensive and systematic approach, the review will follow the guidelines of Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA), involving a structured process of literature search, study selection, data extraction, and synthesis.
The discovery of ecDNA long predates the completion of the Human Genome Project. The term “ecDNA” is used to describe extra-chromosomal DNA found in cancer cells, with sizes reaching hundreds of KB. As early as 1965, Cox et al. reported the discovery of extrachromosomal DNA molecules in human tumor specimens, which often occur in pairs and are therefore called double-minutes. Since then, several studies have found that double microsomes can carry oncogenes, including EGFR and c-Myc, and so on
Recently, Zhang et al. combined rolling cycle amplification and the third generation of Oxford Nanopore sequencing technology to figure out that eccDNA is derived from genomic fragments, including single-fragment self-looping and multi-fragment linked looping. Comparing the eccDNA content of normal cultured cells and apoptotic cells, it was found that the amount of eccDNA in apoptotic cells was significantly increased, indicating that apoptosis can induce the occurrence of eccDNA.
The cyclization of linear DNA fragments is dependent on DNA ligase. There are three common DNA ligase genes in eukaryotes including Lig1, Lig3, and Lig4. When Lig1 and Lig4 are depleted, there is no observable impact on eccDNA production. In typical immune cells, BMDC (bone marrow-derived dendritic cells) and BMDM (bone marrow-derived macrophages), compared with linear genomic DNA fragments of the same size, their eccDNA can induce higher expression of cytokines such as type I interferon (IFN-α, β), interleukin-6 (IL-6), tumor necrosis factor (TNF-α), etc., showing a superior ability to stimulate immune responses.
Amplicon Architect is a tool that uses whole-genome sequencing data to reconstruct ecDNA amplicon structures. The principle of Amplicon Architect is based on two features of ecDNA:
Circle-map takes as input the alignment of the read sequence to the reference genome, which is used by Circle-Map to detect the splitting of the read into two parts to detect genomic rearrangements supporting the circular DNA structure.
ECCsplorer is a bioinformatics pipeline for the automatic detection of eccDNA from double-end sequencing data of amplified circular DNA. ECCsplorer primarily operates through two distinct steps:
Compared with Circle-Map and Amplicon Architect, this process exhibits a higher level of integration and better presentation of results.
CCDA-seq was used to observe the diversity of ecDNA in open chromatin regions, localize the distribution of nucleosomes at the kilobase length scale, and quantify the chromatin state correlation of distal regulatory elements at single-molecule resolution by using m6A MTase methyltransferase to treat genomic DNA, enabling the acquisition of m6A DNA methylation modifications of open chromatin regions at the single-molecule level.
In 2019, Hung et al. found that clusters of approximately 10–100 ecDNAs contributed to intermolecular enhancer-gene interactions in the nucleus and overexpression of proto-oncogenes. EcDNA encoding a variety of different proto-oncogenes forms hubs in a variety of cancer cell types and primary tumors. When clustered with other ecDNA, each ecDNA is more likely to transcribe an oncogene.
Hung et al. identified that ecDNA hubs act as enhanced subsets of the combination,
demonstrating their regulatory function in the transcription of oncogenes. Analysis of the chromosomal targets of ecDNAs at single-molecule resolution revealed that actively expressed oncogenes were spatially clustered in ecDNA-mediated interaction networks.
Chang et al. found that ecDNA aggregated in interphase nuclei can act in clusters, driving intermolecular enhancer signals to amplify oncogene expression. Burgeoning studies have also demonstrated an increase in the frequency of ecDNA between Barrett’s esophagus-associated early esophageal adenocarcinoma (24 %) and stage esophageal adenocarcinoma (43 %), indicating that ecDNA formation occurs during the progression of cancer.
Circular DNA exhibits the ability to undergo chimerism, reintegrating into the linear genome, and promoting the remodeling of oncogenes. study shows that tumors become diverse or heterogeneous when oncogenes are amplified in ecDNA, which allows tumors to complete and maintain high levels of oncogene expression.
Chromosome fragmentation accelerates the rearrangement and amplification of genomic DNA into extrachromosomal DNA, allowing cells to rapidly acquire resistance to changing growth conditions and thus confer resistance to cancer therapy.
A study shows that the glioblastoma cells can develop resistance to the epidermal growth factor receptor (EGFR) -targeting drug erlotinib by eliminating an extrachromosomal copy of the mutated EGFR gene.
EccDNA can be used as a potential biomarker for many diseases related to mutations and genomic rearrangements. EcDNA present in the blood of cancer patients could be isolated and identified by liquid biopsy using Circa-seq. The extrachromosomal circular DNA (ecDNA) can serve as a promising therapeutic target and non-invasive biomarker for prenatal diagnosis, early detection, prognosis, and treatment of gynecological malignancies.
Although the origin of ecDNA has been preliminarily understood, its specific mechanism still needs to be further studied. EcDNA’s unique capacity to mediate ultra-long-distance interactions extends its functional repertoire, opening doors to a myriad of biological functions that warrant exploration. The propensity of ecDNA to facilitate chromatin opening provides a strategic foothold for novel therapeutic directions, including the intriguing prospect of targeting ecDNA in the pursuit of refined tumor treatments. There remains a considerable journey ahead to comprehensively unravel its underlying mechanisms of action and to effectively implement it across various clinical domains, especially in the treatment of tumors.