Publications by Adaptive Scientists
Estimating the ratio of CD4 + to CD8 + T cells using high-throughput sequence data. Journal of Immunological Methods. 18 February 2013. Ryan Emerson, Anna Sherwood, Cindy Desmarais, Sachin Malhotra, Deborah Phippard, Harlan Robins
Mature T cells express either CD8 or CD4, defining two physiologically distinct populations of T cells. CD8+ T cells, or Killer T-cells, and CD4+ T cells, or Helper T cells, effect different aspects of T cell mediated adaptive immunity. Currently, determining the ratio of CD4+ to CD8+ T cells requires flow cytometry or immunohistochemistry. The genomic T cell receptor locus is rearranged during T cell maturation, generating a highly variable T cell receptor locus in each mature T cell. As part of thymic maturation, T cells that will become CD4+ versus CD8+ are subjected to different selective pressures. In this study, we apply high-throughput next-generation sequencing to T cells from both a healthy cohort and a cohort with an autoimmune disease (multiple sclerosis) to identify sequence features in the variable CDR3 region of the rearranged T cell receptor gene that distinguish CD4+ from CD8+ T cells. We identify sequence features that differ between CD4+ and CD8+ T cells, including Variable gene usage and CDR3 region length. We implement a likelihood model to estimate relative proportions of CD4+ and CD8+ T cells using these features. Our model accurately estimates the proportion of CD4+ and CD8+ T cell sequences in samples from healthy and diseased immune systems, and simulations indicate that it can be applied to as few as 1000 T cell receptor sequences; we validate this model using in vitro mixtures of T cell sequences, and by comparing the results of our method to flow cytometry using peripheral blood samples. We believe our computational method for determining the CD4:CD8 ratio in T cell samples from sequence data will provide additional useful information for any samples on which high-throughput TCR sequencing is performed, potentially including some solid tumors.
Shaping of Human Germline IgH Repertoires Revealed by Deep Sequencing. The Journal of Immunology. 14 August 2012. Kevin Larimore, Michael W. McCormick, Harlan S. Robins and Philip D. Greenberg
To understand better how selection processes balance the benefits of Ig repertoire diversity with the risks of autoreactivity and nonfunctionality of highly variable IgH CDR3s, we collected millions of rearranged germline IgH CDR3 sequences by deep sequencing of DNA from mature human naive B cells purified from four individuals and analyzed the data with computational methods. Long HCDR3 regions, often components of HIV-neutralizing Abs, appear to derive not only from incorporation of long D genes and insertion of large N regions but also by usage of multiple D gene segments in tandem. However, comparison of productive and out-of-frame IgH rearrangements revealed a selection bias against long HCDR3 loops, suggesting these may be disproportionately either poorly functional or autoreactive. Our data suggest that developmental selection removes HCDR3 loops containing patches of hydrophobicity, which are commonly found in some auto-antibodies, and at least 69% of the initial productive IgH rearrangements are removed from the repertoire during B cell development. Additionally, we have demonstrated the potential utility of this new technology for vaccine development with the identification in all four individuals of related candidate germline IgH precursors of the HIV-neutralizing Ab 4E10.
High-Throughput Sequencing Detects Minimal Residual Disease in Acute T Lymphoblastic Leukemia. Science Translational Medicine. 16 May 2012. David Wu, Anna Sherwood, Jonathan R. Fromm, Stuart S. Winter, Kimberly P. Dunsmore, Mignon L. Loh, Harvey A. Greisman, Daniel E. Sabath, Brent L. Wood and Harlan Robins
High-throughput sequencing (HTS) of lymphoid receptor genes is an emerging technology that can comprehensively assess the diversity of the immune system. Here, we applied HTS to the diagnosis of T-lineage acute lymphoblastic leukemia/lymphoma. Using 43 paired patient samples, we then assessed minimal residual disease (MRD) at day 29 after treatment. The variable regions of TCRB and TCRG were sequenced using an Illumina HiSeq platform after performance of multiplexed polymerase chain reaction, which targeted all potential V-J rearrangement combinations. Pretreatment samples were used to define clonal T cell receptor (TCR) complementarity-determining region 3 (CDR3) sequences, and paired posttreatment samples were evaluated for MRD. Abnormal T lymphoblast identification by multiparametric flow cytometry was concurrently performed for comparison. We found that TCRB and TCRG HTS not only identified clonality at diagnosis in most cases (31 of 43 for TCRB and 27 of 43 for TCRG) but also detected subsequent MRD. As expected, HTS of TCRB and TCRG identified MRD that was not detected by flow cytometry in a subset of cases (25 of 35 HTS compared with 13 of 35, respectively), which highlights the potential of this technology to define lower detection thresholds for MRD that could affect clinical treatment decisions. Thus, next-generation sequencing of lymphoid receptor gene repertoire may improve clinical diagnosis and subsequent MRD monitoring of lymphoproliferative disorders.
Ultra-Sensitive Detection of Rare T Cell Clones. Journal of Immunological Methods. 31 Jan 2012. HS Robins, C Desmarais, J Matthis, R Livingston, J Andriesen, H Reijonen, CS Carlson, G Nepom, C Yee, K Cerosaletti
To test the precision, accuracy, and sensitivity of our immune profiling assay, T cell clones, with one fixed TCR rearrangement are doped into complex blood cell samples, with the doped T cell clones ranging from 10% of the total sample to 0.001% (one cell in 100,000). Results show precision across five orders of magnitude, and accuracy within an overall factor of three across the 100,000 fold dynamic range. Additionally, the assay is shown to be highly repeatable.
Deep Sequencing of the Human TCRγ and TCRβ Repertoires Suggests that TCRβ Rearranges After αβ and γδ T Cell Commitment. Sci Transl Med. 6 July 2011. Sherwood AM, Desmarais C, Livingston RJ, Andriesen J, Haussler M, Carlson CS, Robins HS
Our data suggest that TCRγ rearranges in all T cells before T lineage commitment, however, rearrangement of the TCRβ locus appears to be restricted after T cell precursors commit to the αβ T lineage. In T cell leukemias and lymphomas, TCRγ is almost always rearranged, and TCRβ is only rearranged in a subset, suggesting the target for monitoring minimal residual disease should be TCRγ.
Overlap and Effective Size of the Human CD8+ T Cell Receptor Repertoire. Sci Transl Med. 1 Sept 2010. Robins HS, Srivastava SK, Campregher PV, Turtle CJ, Andriesen J, Riddell SR, Carlson CS, Warren EH
We sequenced the CDR3 regions from millions of TCRβ chain genes in naïve and memory CD8+ T cells of 7 adults. The CDR3 repertoire in each individual is strongly biased toward specific Vβ-Jβ pair utilization, dominated by sequences containing few inserted nucleotides, and drawn from a defined subset comprising less than 0.1% of the estimated 5 × 1011 possible sequences. The overlap in the naïve CD8+ CDR3 sequence repertoires of any 2 of the individuals is ~7000-fold larger than predicted and appears to be independent of the degree of HLA matching.
Comprehensive assessment of T-cell receptor β-chain diversity in αβ T cells. Blood. 5 Nov 2009. Robins HS, Campregher PV, Srivastava SK, Wacher A, Turtle CJ, Kahsai O, Riddell SR, Warren EH, Carlson CS.
We determine the CDR3 sequence in millions of rearranged TCRβ genes of 2 adults. We find that total TCRβ receptor diversity is at least 4-fold higher than previous estimates, and the diversity in the subset of CD45RO(+) antigen-experienced αβ T cells is at least 10-fold higher than previous estimates. These methods should prove valuable for assessment of αβ T-cell repertoire diversity after hematopoietic cell transplantation, in congenital or acquired immunodeficiency, and during normal aging.
Publications by Adaptive Customers
Plos One 2013
ATM Influences the Efficiency of TCRB Rearrangement, Subsequent TCRB-Dependent T Cell Development, and Generation of the Pre-Selection TCRB CDR3 Repertoire
Hathcock KS1, Bowen S1, Livak F2, Hodes RJ1.
Experimental Immunology Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD1; Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, MD2
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J. Immunol. 2013
The Restricted DH Gene Reading Frame Usage in the Expressed Human Antibody Repertoire Is Selected Based upon its Amino Acid Content.
Benichou J1, Glanville J2, Prak ET3, Azran R4, Kuo TC2, Pons J2, Desmarais C5, Tsaban L4, Louzoun Y4.
Mina and Everard Goodman Faculty of Life Sciences, Bar Illan University, Ramat Fan, Israel1; Protein Engineering and Applied Quantitative Genotherapeutics, Rinat-Pfizer, South San Francisco, CA2; Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA3; Department of Mathematics and Gonda Brain Research Center, Bar Illan University, Ramat Gan, Israel4; and Adaptive Biotechnologies, Seattle, WA5
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Biol. Blood and Bone Marrow Transplant. 2012
Immune Recovery in Adult Patients Following Myeloablative Dual Umbilical Cord Blood, Matched Sibling, and Matched Unrelated Donor Hematopoietic Cell Transplantation
Kanda J, Chiou LW, Szabolcs P, Sempowski GD, Rizzieri DA, Long GD, Sullivan KM, Gasparetto C, Chute JP, Morris A, McPherson J, Hale J, Livingston JA, Broadwater G, Niedzwiecki D, Chao NJ, Horwitz ME.
Division of Cellular Therapy, Department of Medicine, Duke University Medical Center, Durham, North Carolina, USA.
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JCI. 2012
Thymocyte responsiveness to endogenous glucocorticoids is required for immunological fitness.
Mittelstadt PR, Monteiro JP, Ashwell JD.
Center for Cancer Research, NCI and NIAID, NIH, Bethesda, Maryland, USA.
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J. Virol. 2013
High Frequency of Herpesvirus-Specific Clonotypes in the Human T cell Repertoire Can Remain Stable over Decades with Minimal Turnover
Neller MA1, Burrows JM1, Rist MJ1,2, Miles JJ1,2,3, Burrows SR1,2.
Australian Centre for Vaccine Development, Queensland Institute of Medical Research, Brisbane, Australia1; The University of Queensland, Brisbane, Australia2; and Institute of Infection and Immunity, Cardiff University School of Medicine, Cardiff, UK 3
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Mol. Ther. 2013
Allelic Exclusion and Peripheral Reconstitution by TCR Transgenic T cells Arising from Transduced Human Hematopoietic Stem/Progenitor Cells
Giannoni F, Hardee CL, Wherley J, Gschweng E, Senadheera S, Kaufman ML, Chan R, Bahner I, Gersuk V, Wang X, Gjertson D, Baltimore D, Witte ON, Economou JS, Ribas A,
Kohn DB.
Department of Microbiology, Immunology and Molecular Genetics, University of California, Los Angeles, California, USA.
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Biol. Blood Marrow Trasplant. 2013
Fractal Organization of the Human T Cell Repertoire in Health and After Stem Cell Transplantation
Meier J, Roberts C, Avent K, Hazlett A, Berrie J, Payne K, Hamm D, Desmarais C, Sanders C, Hogan KT, Archer KJ, Manjili MH, Toor AA.
Bone Marrow Transplant Program, Department of Internal Medicine, Virginia Commonwealth University, Richmond, Virginia.
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Science Translational Medicine. 2013
CD19-Targeted T Cells Rapidly Induce Molecular Remissions in Adults with Chemotherapy-Refractory Acute Lymphoblastic Leukemia
Brentjens R 1,2,3, Davila M 1, Riviere I 1,2,3,4, Park J 1, Wang X 3,4, Cowell LG 5, Bartido S 4, Stefanski J 4, Taylor C 4, Olszewska M 4, Borquez-Ojeda O 4, Qu J 4, Wasielewska T 4, He Q 4, Bernal Y 1, Rijo IV 6, Hedvat C 6, Kobos R 7, Curran K 7, Steinherz P 7, Jurcic J 1, Rosenblat T 1, Maslak P 1, Frattini M 1, Sadelain M 1,2,3
Department of Medicine, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA 1; Center for Cell Engineering, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA 2; Molecular Pharmacology and Chemistry Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA 3; Cell Therapy and Cell Engineering Facility, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA 4; Department of Clinical Sciences, UT Southwestern, Dallas, TX 75390, USA 5; Department of Pathology, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA 6; Department of Pediatrics, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA 7.
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