Extrachromosomal circular DNA in colorectal cancer: biogenesis, function and potential as therapeutic target

Abstract

Extrachromosomal circular DNA (ecDNA) has gained renewed interest since its discovery more than half a century ago, emerging as critical driver of tumor evolution. ecDNA is highly prevalent in many types of cancers, including colorectal cancer (CRC), which is one of the most deadly cancers worldwide. ecDNAs play an essential role in regulating oncogene expression, intratumor heterogeneity, and resistance to therapy independently of canonical chromosomal alterations in CRC. Furthermore, the existence of ecDNAs is attributed to the patient's prognosis, since ecDNA-based oncogene amplification adversely affects clinical outcomes. Recent understanding of ecDNA put an extra layer of complexity in the pathogenesis of CRC. In this review, we will discuss the current understanding on mechanisms of biogenesis, and distinctive features of ecDNA in CRC. In addition, we will examine how ecDNAs mediate oncogene overexpression, gene regulation, and topological interactions with active chromatin, which facilitates genetic heterogeneity, accelerates CRC malignancy, and enhances rapid adaptation to therapy resistance. Finally, we will discuss the potential diagnostic and therapeutic implications of ecDNAs in CRC.

Fig. 1. Biogenesis, amplification and distribution of ecDNA.

A Formation of ecDNA. (I) Breakage-fusion-bridge (BFB) cycles. Loss of telomere because of genome instability, and the end of the missing telomere fuse with each other to form a chromosomal structure with two centromeres and a dicentric anaphase bridge. The fusion bridge is broken in the late stage of mitosis, keep the genes amplified and circularizing into ecDNA. (II) Chromothripsis model. When chromosomes are catastrophically broken, the DNA double-strand break into some DNA segments, which are randomly linked and cycled to form ecDNA during subsequent DNA repair. (III) Translocation-excision-deletion-amplification (TEDA) model. Segments between chromosomes translocation, DNA fragments between translocation breakpoints are prone to amplification, retention or deletion, and the deleted part is cyclized outside the chromosomes to form ecDNA. (IV) Episome model. Through the way of DNA slippage and R-loop, chromosomes form episomes during genetic recombination, ecDNA generated by cleavage and ligation. B Amplification of ecDNA replicates by rolling circle amplification. C ecDNA distributions. ecDNA can be subject to further clonal evolution, reintegrated into chromosomes, combined with other ecDNAs or eliminated by being trapped inside micronucleus.

Fig. 2. Functions of ecDNA for oncogenesis.

A Transcription. ecDNA can transcribing into RNAs for protein translation or regulating gene expression. B Detection maker. Tumor cells release ecDNA into the blood circulation and thus serve as a tumor detection marker. C Copy-number amplification. Unequal division of ecDNA leads to rapid amplification of oncogenes carried on ecDNA compared to chromosomal DNA. D Intratumoral heterogeneity. Random assignment during ecDNA replication leads to tumor heterogeneity. E Drug resistance. Continuous amplification of ecDNA containing resistance genes leads to drug resistance in tumor cells. F Reshaping cancer genome. If ecDNA is integrated into the upstream of the proto-oncogene, it can enhance the expression of the proto-oncogene, and if integrated into the tumor suppressor gene, it will cause loss of tumor suppressor gene function. G Regulatory circuitry on ecDNA. Multiple ecDNA aggregates to form ecDNA hubs, the enhancers and promoters carried on ecDNA act on protein-coding genes, facilitating the transcription of oncogenes.

Fig. 3. Target ecDNA for colorectal cancer therapy.

A Targeting at biogenesis of ecDNA. The inhibitors of TP53, DNA-PKcs, and TOPII can repress ecDNA formation by inhibiting homologous recombination (HR), nonhomologous terminal ligation (NHEJ), and DNA supercoiling respectively. B Targeting at ecDNA replication and distribution. DNA helicase inhibits rolling circle amplification to suppress the replication process of ecDNA, 5-FU and MTX affect ecDNA replication by directly destroying ecDNA. C Targeting at ecDNA hub. BRD4 inhibitors disrupt aggregation of ecDNA hubs, thus disturbing ecDNA intermolecular regulation and interactions between ecDNA and genomic DNA. D Targeting at elimination of ecDNA. Radiotherapy and HU can reduce ecDNA frequency by promoting the formation of micronuclei, and PARP inhibitors eliminate ecDNA by promoting ecDNA fusion.