Peptide/MHC Tetramer–Based Sorting of CD8+ T Cells to a Leukemia Antigen Yields Clonotypes Drawn Nonspecifically from an Underlying Restricted Repertoire

Hunsucker, Sally A.; McGary, Colleen S.; Vincent, Benjamin G.; Enyenihi, Atim A.; Waugh, Jennifer P.; McKinnon, Karen P.; Bixby, Lisa M.; Ropp, Patricia A.; Coghill, James M.; Wood, William A.; Gabriel, Don A.; Sarantopoulos, Stefanie; Shea, Thomas C.; Serody, Jonathan S.; Alatrash, Gheath; Rodriguez-Cruz, Tania; Lizee, Gregory; Buntzman, Adam; Frelinger, Jeffrey A.; Glish, Gary L.; Armistead, Paul M.

doi:10.1158/2326-6066.cir-14-0001

“…The model predicts the emergence of a limited number of dominant T cell clones, symbolizing antigen driven proliferation and a large number of low frequency clones representing a reservoir of T cell clones which are likely sustained through homeostatic mechanisms. [37,38,39] Recent observation of similar T cell receptor VDJ expressing clones dominating in different patients with a similar HLA type also lends credence to this model and its underlying premise. [40] Incorporating competition, with high affinity antigen directed T cell clones dominating lower affinity antigen directed clones (Supplementary Figure 3B SCT.…”

Section: Discussionmentioning

confidence: 71%

“…This hypothesis is supported by the earlier observation that patients who have oligoclonal T cell recovery following SCT may be more likely to be susceptible to alloreactivity. [37,44] Further support for the notion of oligoclonal T cell growth in response to high affinity and abundant minor histocompatibility antigens comes from the observation of clonal growth in mixed lymphocyte reactions (MLR) predicting loss of renal allografts [45]. MLR has also been used to study, alloreactive T cell clonal populations following SCT and has demonstrated the presence of a large number of dominant and low frequency clones over time.…”

Section: Discussionmentioning

confidence: 99%

Dynamical System Modeling to Simulate Donor T Cell Response to Whole Exome Sequencing-Derived Recipient Peptides Demonstrates Different Alloreactivity Potential In HLA-Matched and Mismatched Donor-Recipient Pairs

Razzaq

¹

,

Scalora

²

,

Kopardé

³

et al. 2015

Preprint

0

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Immune reconstitution kinetics and subsequent clinical outcomes in HLA-matched recipients of allogeneic stem cell transplantation (SCT) are variable and difficult to predict. Considering SCT as a dynamical system, may allow sequence differences across the exomes of the transplant donors and recipients to be used to simulate an alloreactive T cell response, which may allow better clinical outcome prediction. To accomplish this, whole exome sequencing was performed on 34 HLA matched SCT donor-recipient pairs (DRP), and the nucleotide sequence differences translated to peptides. The binding affinity of the peptides to the relevant HLA in each DRP was determined. The resulting array of peptide-HLA binding affinity values in each patient was considered as an operator modifying a hypothetical T cell repertoire vector, in which each T cell clone proliferates in accordance to the logistic equation of growth. Using an iterating system of matrices, each simulated T cell clone's growth was calculated with the steady state population being proportional to the magnitude of the binding affinity of the driving HLA-peptide complex. Incorporating competition between T cell clones responding to different HLA-peptide complexes reproduces a number of features of clinically observed T cell clonal repertoire in the simulated repertoire. These include, sigmoidal growth kinetics of individual T cell clones and overall repertoire, Power Law clonal frequency distribution, increase in repertoire complexity over time with increasing clonal diversity and finally, alteration of clonal dominance when a different antigen array is encountered, such as in stem cell transplantation. The simulated, alloreactive T cell repertoire was markedly different in HLA matched DRP. The patterns were differentiated by rate of growth, and steady state magnitude of the simulated T cell repertoire and demonstrate a possible correlation with survival. In conclusion, exome wide sequence differences in DRP may

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“…Further validation of the model simulating normal physiology comes from the resulting steady state T cell repertoire approximating a Power Law distribution of the T cell clonal frequency for the entire repertoire [18, 19, 20]. The model predicts the emergence of a limited number of dominant T cell clones, symbolizing antigen driven proliferation and a large number of low frequency clones representing a reservoir of T cell clones, which are likely maintained through homeostatic mechanisms [29, 30, 31]. Recent observations of similar T cell receptor VDJ expressing clones dominating in different patients with a similar HLA type also lends credence to this model and its underlying premise [32].…”

Section: Discussionmentioning

confidence: 99%

Dynamical System Modeling to Simulate Donor T Cell Response to Whole Exome Sequencing-Derived Recipient Peptides Demonstrates Different Alloreactivity Potential in HLA-Matched and -Mismatched Donor–Recipient Pairs

Razzaq

¹

,

Scalora

²

,

Kopardé

³

et al. 2016

Biology of Blood and Marrow Transplantation

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Immune reconstitution kinetics and subsequent clinical outcomes in HLA-matched recipients of allogeneic stem cell transplantation (SCT) are variable and difficult to predict. Considering SCT as a dynamical system, may allow sequence differences across the exomes of the transplant donors and recipients to be used to simulate an alloreactive T cell response, which may allow better clinical outcome prediction. To accomplish this, whole exome sequencing was performed on 34 HLA matched SCT donor-recipient pairs (DRP), and the nucleotide sequence differences translated to peptides. The binding affinity of the peptides to the relevant HLA in each DRP was determined. The resulting array of peptide-HLA binding affinity values in each patient was considered as an operator modifying a hypothetical T cell repertoire vector, in which each T cell clone proliferates in accordance to the logistic equation of growth. Using an iterating system of matrices, each simulated T cell clone's growth was calculated with the steady state population being proportional to the magnitude of the binding affinity of the driving HLA-peptide complex. Incorporating competition between T cell clones responding to different HLA-peptide complexes reproduces a number of features of clinically observed T cell clonal repertoire in the simulated repertoire. These include, sigmoidal growth kinetics of individual T cell clones and overall repertoire, Power Law clonal frequency distribution, increase in repertoire complexity over time with increasing clonal diversity and finally, alteration of clonal dominance when a different antigen array is encountered, such as in stem cell transplantation. The simulated, alloreactive T cell repertoire was markedly different in HLA matched DRP. The patterns were differentiated by rate of growth, and steady state magnitude of the simulated T cell repertoire and demonstrate a possible correlation with survival. In conclusion, exome wide sequence differences in DRP may allow simulation of donor alloreactive T cell response to recipient antigens and may provide a quantitative basis for refining donor selection and titration of immunosuppression following SCT.

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“…The TCRβ-chain rearrangement of this clone was analyzed using a modification of another multiplexed RT-PCR protocol for single cell TCRβ analysis. 42 …”

Section: Methodsmentioning

confidence: 99%

Identification and isolation of antigen-specific cytotoxic T lymphocytes with an automated microraft sorting system

Attayek

¹

,

Hunsucker

²

,

Sims

³

et al. 2016

Self Cite

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The simultaneous measurement of T cell function with recovery of individual T cells would greatly facilitate characterizing antigen-specific responses both in vivo and in model systems. We have developed a microraft array methodology that automatically measures the ability of individual T cells to kill a population of target cells and viably sorts specific cells into a 96-well plate for expansion. A human T cell culture was generated against the influenza M1p antigen. Individual microrafts on a 70 × 70 array were loaded with on average 1 CD8+ cell from the culture and a population of M1p presenting target cells. Target cell killing, measured by fluorescence microscopy, was quantified in each microraft. The rates of target cell death among the individual CD8+ T cells varied greatly; however, individual T cells maintained their rates of cytotoxicity throughout the time course of the experiment enabling rapid identification of highly cytotoxic CD8+ T cells. Microrafts with highly active CD8+ T cells were individually transferred to wells of a 96-well plate, using a needle-release device coupled to the microscope. Three sorted T cells clonally expanded. All of these expressed high-avidity T cell receptors for M1p/HLA*02:01 tetramers, and 2 of the 3 receptors were sequenced. While this study investigated single T cell cytotoxicity rates against simple targets with subsequent cell sorting, future studies will involve measuring T cell mediated cytotoxicity in more complex cellular environments, enlarging the arrays to identify very rare antigen specific T cells, and measuring single cell CD4+ and CD8+ T cell proliferation.

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Peptide/MHC Tetramer–Based Sorting of CD8+ T Cells to a Leukemia Antigen Yields Clonotypes Drawn Nonspecifically from an Underlying Restricted Repertoire

Cited by 16 publications

References 26 publications

Dynamical System Modeling to Simulate Donor T Cell Response to Whole Exome Sequencing-Derived Recipient Peptides Demonstrates Different Alloreactivity Potential In HLA-Matched and Mismatched Donor-Recipient Pairs

Dynamical System Modeling to Simulate Donor T Cell Response to Whole Exome Sequencing-Derived Recipient Peptides Demonstrates Different Alloreactivity Potential In HLA-Matched and Mismatched Donor-Recipient Pairs

Dynamical System Modeling to Simulate Donor T Cell Response to Whole Exome Sequencing-Derived Recipient Peptides Demonstrates Different Alloreactivity Potential in HLA-Matched and -Mismatched Donor–Recipient Pairs

Identification and isolation of antigen-specific cytotoxic T lymphocytes with an automated microraft sorting system

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