Abstract 993: Whole exome sequencing of CTCs as a window into metastatic cancer
Authors: Adalsteinsson Viktor A. , Lohr Jens G. , Cibulskis Kristian , Choudhury Atish D. , Rosenberg Mara , Cruz-Gordillo Peter , Francis Joshua , Zhang ChengZhong , Shalek Alexander K. , Satija Rahul , Trombetta John T. , Lu Diana , Tallapragada Naren , Tahirova Narmin T. , Kim Sora , Blumenstiel Brendan , Sougnez Carrie , Auclair Daniel , Allen Eliezer M. Van , Nakabayashi Mari , Lis Rosina T. , Lee Gwo-Shu M. , Li Tiantian , Chabot Matthew S. , Taplin Mary-Ellen , Clancy Thomas E. , Loda Massimo , Regev Aviv , Meyerson Matthew , Hahn William C. , Kantoff Philip W. , Golub Todd R. , Getz Gad , Boehm Jesse S. , Love J Christopher 
Published: May 2015

Abstract
Comprehensive analysis of cancer genomes in clinical settings holds the promise to inform prognoses and guide the deployment of precise cancer treatments. A major barrier, however, is the inaccessibility of adequate metastatic tissue for accurate genomic analysis. The recognition that circulating tumor cells (CTCs) are present in many advanced cancer patients suggests an exciting opportunity to overcome this challenge. For instance, if CTCs could be comprehensively sequenced, it would be possible to obtain an orthogonal sample of the tumor burden_including subsets of transiting cells bound for metastatic colonization_potentially yielding new insights to complement the static sampling of resected or biopsied lesions.
We report an integrated process to isolate, qualify, and sequence whole exomes of CTCs with high fidelity, using a census-based sequencing strategy. We isolated CTCs by magnetic bead purification (Illumina MagSweeper) from the blood of patients with prostate cancer, and integrated a nanowell platform to automatically image and recover candidate single CTCs. We then developed a strategy to qualify individual CTC-derived libraries for DNA sequencing after whole genome amplification, and established an analytical framework for accurate calling of mutations using census-based sequencing and MuTect. Whole exome sequencing was performed on 20 single CTCs, obtained from a patient with advanced prostate cancer. We validated our sequencing process by comparing CTC-derived mutations to mutations found in a lymph node metastasis and nine separate cores of the primary tumor. 51 of 73 CTC mutations (70%) were observed in the metastasis or the primary tumor. Moreover, we identified 9 early trunk mutations and 56 metastatic trunk mutations in the non-CTC tumor samples and found 100% and 73% of these, respectively, in CTC exomes. Our work demonstrates the feasibility of CTC sequencing and the ability to confidently call somatic mutations. CTCs may therefore represent a non-invasive window into the mutational landscape of metastatic cancer, and may have utility for genomics in clinical practice.
Citation Format: Viktor A. Adalsteinsson, Jens G. Lohr, Kristian Cibulskis, Atish D. Choudhury, Mara Rosenberg, Peter Cruz-Gordillo, Joshua Francis, ChengZhong Zhang, Alexander K. Shalek, Rahul Satija, John T. Trombetta, Diana Lu, Naren Tallapragada, Narmin T. Tahirova, Sora Kim, Brendan Blumenstiel, Carrie Sougnez, Daniel Auclair, Eliezer M. Van Allen, Mari Nakabayashi, Rosina T. Lis, Gwo-Shu M. Lee, Tiantian Li, Matthew S. Chabot, Mary-Ellen Taplin, Thomas E. Clancy, Massimo Loda, Aviv Regev, Matthew Meyerson, William C. Hahn, Philip W. Kantoff, Todd R. Golub, Gad Getz, Jesse S. Boehm, J Christopher Love. Whole exome sequencing of CTCs as a window into metastatic cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 993. doi:10.1158/1538-7445.AM2014-993