Abstract 4017: Dissecting the clonal hierarchy of cancer-driving genomic lesions.
Authors: Prandi Davide , Baca Sylvan C. , Lawrence Michael S. , Mosquera Juan Miguel , Romanel Alessandro , Drier Yotam , Park Kyung , Kitabayashi Naoki , MacDonald Theresa Y. , Van Allen Eliezer , Kryukov Gregory V. , Theurillat Jean-Philippe , Soong T. David , Nickerson Elizabeth , Auclair Daniel , Tewari Ashutosh , Beltran Himisha , Onofrio Robert C. , Boysen Gunther , Guiducci Candace , Barbieri Christopher E. , Cibulskis Kristian , Sivachenko Andrey , Carter Scott L. , Saksena Gordon , Voet Douglas , Ramos Alex H. , Winckler Wendy , Cipicchio Michelle , Ardlie Kristin , Kantoff Philip W. , Berger Michael F. , Gabriel Stacey B. , Golub Todd R. , Meyerson Matthew , Lander Eric S. , Elemento Olivier , Getz Gad , Demichelis Francesca , Rubin Mark A. , Garraway Levi A. 
Published: August 2014

Characterizing the genomic evolution of cancer is critical to understanding disease progression and identifying potential therapeutic targets. By examining the clonal hierarchy of genomic lesions in common tumors, it would be possible to reconstruct the path of oncogenic events that drive carcinogenesis. Reliable assessment of such paths from high-throughput genome sequencing data is complicated by the admixture of normal DNA in tumor samples and by reduced data signal for highly subclonal events. We introduce an approach that exploits individuals’ genetic background by using the abundant germline SNP genotype data provided by whole genome sequence coverage to assess the clonality of genomic alterations, including copy number changes, rearrangements, and point mutations.

We developed a novel algorithm, CLONET (CLONality Estimate in Tumors), which analyzes patient-specific heterozygous SNP loci (informative SNPs) and mono-allelic somatic deletions to assess levels of stromal DNA admixture and infer the clonal status of each aberration. For every mono allelic deletion, CLONET assesses the allelic fractions of informative SNPs to determine the apparent proportion of normal cells DNA. Next, through a conservative use of simulation-based error estimates, deletions with the lowest proportions of normal DNA reads are considered clonal. For point mutations, the tumor allelic fraction is corrected for stromal DNA admixture level and subclonality is inferred when it differs significantly from the expected value for clonal lesions. Similarly, the proportions of reads that span each side of a putative breakpoint involved in a rearrangement are matched against the expected values. CLONET also addresses tumor aneuploidy by searching for chromosomes with coverage and allelic fractions of informative SNPs not consistent with a diploid genome.

CLONET was tested on 55 whole genome sequences from prostate cancers, a highly heterogeneous tumor type, to catalogue the accumulation of somatic alterations during oncogenesis and progression. In 98% of the cases CLONET made confident assessment of admixture and clonality. We observed consistent clonal lesions involving NKX3-1, the 3Mb region between TMPRSS2 and ERG and FOXP1, as well as early point mutations in SPOP and FOXA1. Overall, we observed a higher rate of subclonal protein-coding point mutation versus deletions (p-value &lt; 10−7). We validated this approach by IHC and FISH for predicted clonal and sub-clonal events. A predicted subclonal homozygous deletion of CHD1 was confirmed by FISH that demonstrated the presence of both nuclei with homozygous and with hemizygous deletion of CHD1. Finally, to assess the general validity of CLONET, we analyzed data from 53 additional tumor genomes, including 25 melanomas and 28 lung adenocarcinomas.

In summary, our results imply the existence of consensus paths of tumor carcinogenesis that favor dysregulation of cancer genes in a defined sequence.

Citation Format: Davide Prandi, Sylvan C. Baca, Michael S. Lawrence, Juan Miguel Mosquera, Alessandro Romanel, Yotam Drier, Kyung Park, Naoki Kitabayashi, Theresa Y. MacDonald, Eliezer Van Allen, Gregory V. Kryukov, Jean-Philippe Theurillat, T. David Soong, Elizabeth Nickerson, Daniel Auclair, Ashutosh Tewari, Himisha Beltran, Robert C. Onofrio, Gunther Boysen, Candace Guiducci, Christopher E. Barbieri, Kristian Cibulskis, Andrey Sivachenko, Scott L. Carter, Gordon Saksena, Douglas Voet, Alex H. Ramos, Wendy Winckler, Michelle Cipicchio, Kristin Ardlie, Philip W. Kantoff, Michael F. Berger, Stacey B. Gabriel, Todd R. Golub, Matthew Meyerson, Eric S. Lander, Olivier Elemento, Gad Getz, Francesca Demichelis, Mark A. Rubin, Levi A. Garraway. Dissecting the clonal hierarchy of cancer-driving genomic lesions. [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 4017. doi:10.1158/1538-7445.AM2013-4017