Viktor Adalsteinsson, PhD

Group Leader, Broad Institute

Selected Publications

• V.A. Adalsteinsson, G. Ha, S.S. Freeman, A.D. Choudhury, D.G. Stover, H.A. Parsons, G. Gydush, S.C. Reed, D. Loginov, D. Livitz, D. Rosebrock, I. Leshchiner, J. Kim, C. Stewart, M. Rosenberg, J.M. Francis, C.Z. Zhang, O. Cohen, C. Oh, H. Ding, P. Polak, M. Lloyd, S. Mahmud, K. Helvie, M.S. Merrill, R.A. Santiago, E.P. O'Connor, S.H. Jeong, R. Leeson, R.M. Barry, J.F. Kramkowski, Z. Zhang, L. Polacek, J.G. Lohr, N.M. Oliver, L. Marini, A.G. Waks, L.C. Harshman, S.M. Tolaney, E.M. Van Allen, E.P. Winer, N.U. Lin, M. Nakabayashi, M.E. Taplin, C.M. Johannessen, L.A. Garraway, T.R. Golub, J.S. Boehm, N. Wagle, G. Getz, J.C. Love, M. Meyerson, Scalable approach for whole-exome sequencing of cell-free DNA from patients with metastatic cancer, submitted 
• Abstract: Whole-exome sequencing of cell-free DNA (cfDNA) stands to enable comprehensive profiling of tumors from blood. The ability to apply whole-exome sequencing of cfDNA in clinical settings, however, is dependent upon the establishment of scalable approaches and rigorous comparisons of cfDNA and tumor whole-exomes. Here, we describe a systematic approach that uses ultra-low-pass whole-genome sequencing (ULP-WGS, 0.1x coverage) to qualify cfDNA for whole-exome sequencing based on tumor content. Whole-exome sequencing of cfDNA and matched tumor biopsies from 23 patients revealed high concordance of clonal somatic mutations (90%), copy number alterations (80%), mutational signatures, and neoantigens. Subsequent screening of 879 blood samples from 333 patients with metastatic prostate or breast cancers revealed that 42% of patients had sufficient tumor content in cfDNA for standard depth of whole-exome sequencing. Our study demonstrates that cfDNA provides an accurate proxy for metastatic tumors and is applicable to many patients.

• J.G. Lohr, V.A. Adalsteinsson, K. Cibulskis, A.D. Choudhury, M. Rosenberg, P. Cruz-Gordillo, J. Francis, C.Z. Zhang, A.K. Shalek, R. Satija, J.T. Trombetta, D. Lu, N. Tallapragada, N. Tahirova, S. Kim, B. Blumenstiel, C. Sougnez, A. Lowe, B. Wong, D. Auclair, E.M. Van Allen, M. Nakabayashi, R.T. Lis, G.S.M. Lee, T. Li, M.S. Chabot, A. Ly, M.E. Taplin, T.E. Clancy, M. Loda, A. Regev, M. Meyerson, W.C. Hahn, P.W. Kantoff, T.R. Golub, G. Getz, J.S. Boehm, J.C. Love, Whole exome sequencing of CTCs as a window into metastatic prostate cancer, Nature Biotechnology, 2014, 32(5), pp 479-484
• Abstract: Comprehensive analyses of cancer genomes promise to inform prognoses and precise cancer treatments. A major barrier, however, is inaccessibility of metastatic tissue. A potential solution is to characterize circulating tumor cells (CTCs), but this requires overcoming the challenges of isolating rare cells and sequencing low-input material. Here we report an integrated process to isolate, qualify and sequence whole exomes of CTCs with high fidelity using a census-based sequencing strategy. Power calculations suggest that mapping of >99.995% of the standard exome is possible in CTCs. We validated our process in two patients with prostate cancer, including one for whom we sequenced CTCs, a lymph node metastasis and nine cores of the primary tumor. Fifty-one of 73 CTC mutations (70%) were present in matched tissue. Moreover, we identified 10 early trunk and 56 metastatic trunk mutations in the non-CTC tumor samples and found 90% and 73% of these mutations, respectively, in CTC exomes. This study establishes a foundation for CTC genomics in the clinic.

• C.Z. Zhang, V.A. Adalsteinsson, J. Francis, J. Jung, C. Maire, H. Cornils, M. Leibowitz, D. Pellman, K.L. Ligon, M. Meyerson, J.C. Love, Predicting complexity and power of single-cell DNA sequencing libraries, Nature Communications, 6(6822), 2015
• Abstract: Artifacts introduced in whole-genome amplification (WGA) make it difficult to derive accurate genomic information from single-cell genomes and require different analytical strategies from bulk genome analysis. Here, we describe statistical methods to quantitatively assess the amplification bias resulting from whole-genome amplification of single-cell genomic DNA. Analysis of single-cell DNA libraries generated by different technologies revealed universal features of the genome coverage bias predominantly generated at the amplicon level (1–10 kb). The magnitude of coverage bias can be accurately calibrated from low-pass sequencing (B0.1􏰁) to predict the depth-of-coverage yield of single-cell DNA libraries sequenced at arbitrary depths. We further provide a benchmark comparison of single-cell libraries generated by multi-strand displacement amplification (MDA) and multiple annealing and looping-based amplification cycles (MALBAC). Finally, we develop statistical models to calibrate allelic bias in single-cell whole-genome amplification and demonstrate a census-based strategy for efficient and accurate variant detection from low-input biopsy samples. 

• J. Francis, C.Z. Zhang, C. Maire, J. Jung, V. Manzo, V.A. Adalsteinsson, H. Homer, S. Haidar, B. Blumensteil, C.S. Pedamallu, A. Ligon, J.C. Love, M. Meyerson, K. Ligon, EGFR variant heterogeneity in glioblastoma resolved through single-nucleus sequencing, Cancer Discovery, 2014, DOI 10.1158/2159-8290.CD-13-0879
• Abstract: Glioblastomas (GBM) with EGFR amplification represent approximately 50% of newly diagnosed cases, and recent studies have revealed frequent coexistence of multiple EGFR aberrations within the same tumor, which has implications for mutation cooperation and treatment resistance. However, bulk tumor sequencing studies cannot resolve the patterns of how the multiple EGFR aberrations coexist with other mutations within single tumor cells. Here, we applied a population-based single-cell whole-genome sequencing methodology to characterize genomic heterogeneity in EGFR-amplified glioblastomas. Our analysis effectively identified clonal events, including a novel translocation of a super enhancer to the TERT promoter, as well as subclonal LOH and multiple EGFR mutational variants within tumors. Correlating the EGFR mutations onto the cellular hierarchy revealed that EGFR truncation variants (EGFRvII and EGFR carboxyl-terminal deletions) identified in the bulk tumor segregate into nonoverlapping subclonal populations. In vitro and in vivo functional studies show that EGFRvII is oncogenic and sensitive to EGFR inhibitors currently in clinical trials. Thus, the association between diverse activating mutations in EGFR and other subclonal mutations within a single tumor supports an intrinsic mechanism for proliferative and clonal diversification with broad implications in resistance to treatment.

• V.A. Adalsteinsson, N. Tahirova, N. Tallapragada, X. Yao, L. Campion, A. Angelini, T.B. Douce, C. Huang, B. Bowman, C.A. Williamson, D.S. Kwon, K.D. Wittrup, J.C. Love, Single cells from human primary colorectal tumors exhibit polyfunctional heterogeneity in secretions of ELR+ CXC chemokines, Integrative Biology, 2013, 5, pp 1272 – 1281
• Abstract: Cancer is an inflammatory disease of tissue that is largely influenced by the interactions between multiple cell types, secreted factors, and signal transduction pathways. While single-cell sequencing continues to refine our understanding of the clonotypic heterogeneity within tumors, the complex interplay between genetic variations and non-genetic factors ultimately affects therapeutic outcome. Much has been learned through bulk studies of secreted factors in the tumor microenvironment, but the secretory behavior of single cells has been largely uncharacterized. Here we directly profiled the secretions of ELR+ CXC chemokines from thousands of single colorectal tumor and stromal cells, using an array of subnanoliter wells and a technique called microengraving to characterize both the rates of secretion of several factors at once and the numbers of cells secreting each chemokine. The ELR+ CXC chemokines are highly redundant, pro-angiogenic cytokines that signal via the CXCR1 and CXCR2 receptors, influencing tumor growth and progression. We find that human primary colorectal tumor and stromal cells exhibit polyfunctional heterogeneity in the combinations and magnitudes of secretions for these chemokines. In cell lines, we observe similar variance: phenotypes observed in bulk can be largely absent among the majority of single cells, and discordances exist between secretory states measured and gene expression for these chemokines among single cells. Together, these measures suggest secretory states among tumor cells are complex and can evolve dynamically. Most importantly, this study reveals new insight into the intratumoral phenotypic heterogeneity of human primary tumors.

• A. Sharei, V.A. Adalsteinsson, A.S. Liss, N. Cho, T. Kamath, S. Jiang, N. Petal, J.K. Park, S.P. Thayer, R. Langer, K.F. Jensen, J.C. Love, Size-selective delivery platform for labeling and manipulation of CTCs., Small, accepted
• Abstract: Identifying and separating a subpopulation of cells from a heterogeneous mixture are essential elements of biological research. Current approaches require detailed knowledge of unique cell surface properties of the target cell population. We describe a method that exploits size differences of cells to facilitate selective intra-cellular delivery using a high throughput microfluidic device. Cells traversing a constriction within this device undergo a transient disruption of the cell membrane that allows for cytoplasmic delivery of cargo. This platform is able to deliver cargo to diverse cell sizes and we show that a 4 micron difference in cell diameter is sufficient to confer unique constriction width preferences for optimum delivery. We demonstrate application of this approach by selectively delivering dextran-conjugated fluorophores to circulating tumor cells in patient blood allowing for their subsequent isolation and genomic characterization.

• X. Yao, A.D. Choudhury, Y.J. Yamanaka, V.A. Adalsteinsson, T.M. Gierahn, C.A. Williamson, C.R. Lamb, M.E. Taplin, M. Nakabayashi, G.S.M. Lee, T. Li, M. Chabot, J.S. Boehm, P.W. Kantoff, W.C. Hahn, K.D. Wittrup, J.C. Love, Functional analysis of single cells identifies a rare subset of circulating tumor cells with malignant traits, Integrative Biology, 2014, 6(4), pp 388 - 398
• Abstract: Ample evidence supports genetic and functional heterogeneity in primary tumors, but it remains unclear whether circulating tumor cells (CTCs) also exhibit the same hierarchical organization. We examined the functional diversity of viable, single CTCs using an array of subnanoliter wells (nanowells). The compartmentalization of single cells by nanowells allowed clonal comparison and mapping of heterogeneity of single cells or preformed clusters of cells. By measuring the short-term viability, invasiveness and secretory profiles of individual CTCs, it was evident that only a rare subset of CTCs possessed malignant traits indicative of metastatic potential in late-stage, progressing metastatic castration-resistant prostate cancer (mCRPC) patients. These CTCs were resistant to anoikis after being in the circulation, were invasive in their epithelial state, or secreted proteases capable of cleaving peptide substrates. Every CTC observed, however, did not exhibit such metastatic potential, suggesting that enumeration of CTCs alone may be insufficient to understand metastasis or stratify patients.

• V.A. Adalsteinsson, O. Parajuli, S. Kepics, A. Gupta, W. Brian Reeves, J. Hahm, Ultrasensitive Detection of Cytokines Enabled by Nanoscale ZnO Arrays, Journal of Analytical Chemistry, 2008, 80 (17), pp 6594–6601
• Abstract: Early detection of disease markers can provide higher diagnostic power and improve disease prognosis. We demonstrate the use of zinc oxide nanorod (ZnO NR) arrays in a straightforward, reliable, and ultrasensitive detection of the cytokines interleukin-18 and tumor necrosis factor-r. Specifically, we exploit the fluorescence- enhancing properties of ZnO NR platforms in cytokine assays involving both a pure buffer and urine. The detection sensitivity achieved using this ZnO NR method is in the subfemtogram per milliliter level, which is 3-4 orders of magnitude more sensitive than conventional assay detection limits. This unparalleled detection sensi- tivity is achieved without the need for indirect enzyme reactions or specialized instrumentation. We highlight various advantages of using ZnO NR arrays in the ultra- sensitive profiling of cytokine levels. Key advantages include robustness of NR arrays, simple and direct assay schemes, high-throughput and multiplexing capabilities, and the ability to correlate directly measured signals to cytokine levels. In conjunction with the extremely high sensitivity demonstrated in this work, our ZnO NR array- based approach may be highly beneficial in early detection of many cytokine-implicated diseases.