Esophageal adenocarcinoma is a prominent example of cancer characterized by frequent amplifications in oncogenes. However, the mechanisms leading to amplicons that involve breakage-fusion-bridge cycles and extrachromosomal DNA are poorly understood. Here, we use 710 esophageal adenocarcinoma cases with matched samples and patient-derived organoids to disentangle complex amplicons and their associated mechanisms. Short-read sequencing identifies ERBB2, MYC, MDM2, and HMGA2 as the most frequent oncogenes amplified in extrachromosomal DNAs. We resolve complex extrachromosomal DNA and breakage-fusion-bridge cycles amplicons by integrating of de-novo assemblies and DNA methylation in nine long-read sequenced cases. Complex amplicons shared between precancerous biopsy and late-stage tumor, an enrichment of putative enhancer elements and mobile element insertions are potential drivers of complex amplicons’ origin. We find that patient-derived organoids recapitulate extrachromosomal DNA observed in the primary tumors and single-cell DNA sequencing capture extrachromosomal DNA-driven clonal dynamics across passages. Prospectively, long-read and single-cell DNA sequencing technologies can lead to better prediction of clonal evolution in esophageal adenocarcinoma.

食管腺癌是癌症的一个突出例子，其特征是癌基因频繁扩增。然而，导致涉及断裂-融合-桥循环和染色体外DNA的扩增子的机制知之甚少。在这里，我们使用 710 例食管腺癌病例以及匹配的样本和患者来源的类器官来解开复杂的扩增子及其相关机制。短读长测序将ERBB2、MYC、MDM2和HMGA2确定为染色体外 DNA 中最常见的扩增癌基因。我们通过在九个长读长测序案例中整合从头组装和 DNA 甲基化来解决复杂的染色体外 DNA 和断裂-融合-桥循环扩增子。癌前活检和晚期肿瘤之间共享的复杂扩增子、推定增强子元件的富集和移动元件插入是复杂扩增子起源的潜在驱动因素。我们发现，源自患者的类器官再现了在原发性肿瘤中观察到的染色体外 DNA，并且单细胞 DNA 测序捕获了染色体外 DNA 驱动的跨代克隆动态。预计，长读长和单细胞 DNA 测序技术可以更好地预测食管腺癌的克隆进化。