293 Background: Genomic characterization of urothelial carcinoma of the bladder (UCB) has begun to reveal significant intertumor heterogeneity when comparing samples from different subjects. As in other malignancies, intratumor heterogeneity, which may allow for tumor evolution and adaption, poses a significant challenge to personalized-medicine strategies. Methods: To examine UCB tumor evolution and heterogeneity, we performed next-generation targeted sequencing on multiple temporally and spatially separated bladder tumors obtained at time of transurethral resection (TUR) and radical cystectomy (RC). Specimens were analyzed using a next-generation, targeted sequencing assay designed to identify point mutations, indels, and copy number alterations in 300 cancer-associated genes. Results: Phylogenetic reconstruction revealed evidence of branched evolutionary growth. Evaluation of multiple tumors from individual subjects identified both shared and unique potential driver mutations. Evidence of convergent phenotypic evolution was detected through analysis of multiple distinct tumors from several subjects. For example, three separate tumors in one subject shared a common PIK3CA mutation (E453K) and had unique second mutations in PIK3CA (E542V, E545K, and E545Q, respectively). In another subject, distinct inactivating mutations of EP300 were identified in two temporally separated tumor samples. Macrodissection of single tumors into non-invasive and invasive components revealed significant intratumor heterogeneity; one case illustrates how analysis of a muscle-invasive TUR specimen could result in undersampling and thereby miss the tumor clone that persisted at time of RC. Conclusions: We demonstrate branched evolution of UCB through genomic analyses of multiple temporally and spatially distinct bladder tumors from individual subjects. Macrodissection of individual tumor samples identified significant intratumor heterogeneity. These concepts may present major challenges to personalized-medicine approaches that rely on sampling of a single tumor at a specific timepoint in the evolution of a patient’s UCB.
Intratumor Heterogeneity and Branched Evolution Revealed by Multiregion Sequencing
