The interplay of mutagenesis and ecDNA shapes urothelial cancer evolution
Duy Nguyen, William F. Hooper, Weisi Liu, Timothy R. Chu, Heather Geiger, Jennifer Shelton, Minita Shah, Zoe R. Goldstein, Lara Winterkorn, Adrienne Helland, Michael Sigouros, Jyothi Manohar, Jenna E. Moyer, Majd Al Assaad, Alissa Semaan, Sandra Cohen, Florencia P. Madorsky Rowdo, David Wilkes, Mohamed Osman, Rahul Singh, Andrea Sboner, Henkel Valentine, Philip H. Abbosh, Scott T. Tagawa, David M. Nanus, Jones T. Nauseef, Cora N. Sternberg, Ana M. Molina, Douglas S. Scherr, Giorgio Inghirami, Juan Miguel Mosquera, Olivier Elemento, Nicolas Robine, Bishoy M. Faltas
Abstract
Advanced urothelial cancer is a frequently lethal disease characterized by marked genetic heterogeneity1. In this study, we investigated the evolution of genomic signatures caused by endogenous and external mutagenic processes and their interplay with complex structural variants (SVs). We superimposed mutational signatures and phylogenetic analyses of matched serial tumours from patients with urothelial cancer to define the evolutionary dynamics of these processes. We show that APOBEC3-induced mutations are clonal and early, whereas chemotherapy induces mutational bursts of hundreds of late subclonal mutations. Using a genome graph computational tool2, we observed frequent high copy-number circular amplicons characteristic of extrachromosomal DNA (ecDNA)-forming SVs. We characterized the distinct temporal patterns of APOBEC3-induced and chemotherapy-induced mutations within ecDNA-forming SVs, gaining new insights into the timing of these mutagenic processes relative to ecDNA biogenesis. We discovered that most CCND1 amplifications in urothelial cancer arise within circular ecDNA-forming SVs. ecDNA-forming SVs persisted and increased in complexity, incorporating additional DNA segments and contributing to the evolution of treatment resistance. Oxford Nanopore Technologies long-read whole-genome sequencing followed by de novo assembly mapped out CCND1 ecDNA structure. Experimental modelling of CCND1 ecDNA confirmed its role as a driver of treatment resistance. Our findings define fundamental mechanisms that drive urothelial cancer evolution and have important therapeutic implications. Whole-genome sequencing of matched serial tumours from patients identifies two key mutagenic factors (APOBEC3 and chemotherapy) and extrachromosomal DNA-forming structural variants that drive treatment resistance in urothelial cancer.