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SUBSTITUTION AND SITE-SPECIFIC SELECTION DRIVING B CELL AFFINITY MATURATION IS CONSISTENT ACROSS INDIVIDUALS
"... ABSTRACT. The antibody repertoire of each individual is continuously updated by the evolutionary process of B cell receptor mutation and selection. It has re-cently become possible to gain detailed information concerning this process through high-throughput sequencing. Here, we develop modern statis ..."
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ABSTRACT. The antibody repertoire of each individual is continuously updated by the evolutionary process of B cell receptor mutation and selection. It has re-cently become possible to gain detailed information concerning this process through high-throughput sequencing. Here, we develop modern statistical molecular evo-lution methods for the analysis of B cell sequence data, and then apply them to a very deep short-read data set of B cell receptors. We find that the substitution pro-cess is conserved across individuals but varies significantly across gene segments. We investigate selection on B cell receptors using a novel method that side-steps the difficulties encountered by previous work in differentiating between selection and motif-driven mutation; this is done through stochastic mapping and empiri-cal Bayes estimators that compare the evolution of in-frame and out-of-frame re-arrangements. We use this new method to derive a per-residue map of selection, which we find is dominated by purifying selection, though not uniformly so.
RESEARCH ARTICLE Detection and Tracking of NY-ESO-1-Specific CD8+ T Cells by High-Throughput T Cell Receptor β (TCRB) Gene Rearrangements Sequencing in a Peptide-Vaccinated Patient
"... Comprehensive immunological evaluation is crucial for monitoring patients undergoing antigen-specific cancer immunotherapy. The identification and quantification of T cell responses is most important for the further development of such therapies. Using well-char-acterized clinical samples from a hig ..."
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Comprehensive immunological evaluation is crucial for monitoring patients undergoing antigen-specific cancer immunotherapy. The identification and quantification of T cell responses is most important for the further development of such therapies. Using well-char-acterized clinical samples from a high responder patient (TK-f01) in an NY-ESO-1f peptide vaccine study, we performed high-throughput T cell receptor β-chain (TCRB) gene next generation sequencing (NGS) to monitor the frequency of NY-ESO-1-specific CD8+ T cells. We compared these results with those of conventional immunological assays, such as IFN-γ capture, tetramer binding and limiting dilution clonality assays. We sequenced human TCRB complementarity-determining region 3 (CDR3) rearrangements of two NY-ESO-1f-specific CD8+ T cell clones, 6-8L and 2F6, as well as PBMCs over the course of peptide vaccination. Clone 6-8L possessed the TCRB CDR3 gene TCRBV11-03*01 and BJ02-01*01 with amino acid sequence CASSLRGNEQFF, whereas 2F6 possessed TCRBV05-08*01 and BJ02-04*01 (CASSLVGTNIQYF). Using these two sequences as models, we evaluated the frequency of NY-ESO-1-specific CD8+ T cells in PBMCs ex vivo. The 6-8L
