Abstract 4603: Intratumor heterogeneity in clear cell renal cell carcinoma (ccRCC): Multi-region sequencing redefines the mutational landscape of ccRCCs.

Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Background: Current efforts to define the genomic landscapes of solid tumours are based on the analysis of single tumour biopsies. We recently demonstrated through multi-region exome sequencing (M-seq) of 2 ccRCC that these cancers are genetically heterogeneous (Gerlinger et al. NEJM, 2012). Thus, single biopsy approaches are likely to underestimate the genetic complexity and the incidence of driver gene mutations. Methods: We applied M-seq and copy number profiling to an average of 7 regions from each of 7 advanced ccRCCs to reveal the genomic landscapes and to reconstructed the life histories of these tumours through phylogenetic analysis. Results: M-seq identified approximately 100 non-synonymous somatic mutations per case on average; twice as many mutations than revealed on average in a single biopsy or the 54 mutation found on average in single biopsies of metastatic ccRCCs analysed by The Cancer Genome Atlas. M-seq revealed that all tumours were heterogeneous with more than 50% of non-synonymous somatic mutations and copy number aberrations not present across all analysed tumour regions. The mutational spectrum and chromosomal instability scores differed between regions of individual tumours. Phylogenetic reconstruction showed branched evolution in all cases. Loss of chromosome 3p and mutations in the VHL and PBRM1 genes were the only ccRCC drivers which were altered ubiquitously throughout all regions in all tumours where they occurred. These were the only known ccRCC driver mutations which were recurrently located on the trunk of the phylogenetic trees, indicating that these truncal genetic events are sufficient for tumour initiation. All other known ccRCC driver mutations and the vast majority of chromosomal driver aberrations were confined to spatially separated subclones, represented as branches on the phylogenetic trees. Thus, genetic alterations of VHL, PBRM1 and chromosome 3p appear to initiate the generation of genetic diversity and the acquisition of additional driver events promoting expansion of spatially separated subclones. The truncal driver mutations were similar across all tumours but clinical outcomes differed significantly between patients, indicating that subclonal drivers may determine clinical outcomes. Conclusions: M-seq revealed intra-tumour heterogeneity and parallel evolution of multiple subclones in each of 7 metastatic tumors, indicating that these are common characteristic of ccRCCs. VHL, PBRM1 and chromosome 3p were the only recurrent truncal driver events identified. All other driver mutations were heterogeneous and these are likely to influence clinical outcome. Thus, M-seq reveals complex heterogeneous genomic landscapes in ccRCCs and demonstrates that the detection of subclonal driver mutations may be highly relevant for personalized cancer medicine approaches. Citation Format: Marco Gerlinger, Stuart Horswell, James Larkin, Andrew J. Rowan, Pierre Martinez, Ignacio Varela, Claudio R. Santos, Rosalie Fisher, Max P. Salm, Zoltan Szallasi, Nicholas Matthews, Aengus Stewart, P Andrew Futreal, Charles Swanton. Intratumor heterogeneity in clear cell renal cell carcinoma (ccRCC): Multi-region sequencing redefines the mutational landscape of ccRCCs. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4603. doi:10.1158/1538-7445.AM2013-4603