generating-de-novo-antigen-specific-human-t-cell-receptors-retroviral

Generating <em>De Novo</em> Antigen-specific Human T Cell Receptors by Retroviral Transduction of Centric Hemichain

T cell receptors (TCRs) are used clinically to direct the specificity of T cells to target tumors as a promising modality of immunotherapy. Therefore, ...

University of Toronto

University Health Network

Recent high throughput sequencing analysis has revealed that the TCRβ repertoire is largely different between CD8(+) and CD4(+) T cells. Here, we show that the transduction of SIG35α, the public chain-centric HLA-A*02:01(A2)/MART127-35 TCRα hemichain, conferred A2/MART127-35 reactivity to a substantial subset of both CD8(+) and CD4(+) T cells regardless of their HLA-A2 positivity. T cells individually reconstituted with SIG35α and different A2/MART127-35 TCRβ genes isolated from CD4(+) or CD8(+) T cells exhibited a wide range of avidity. Surprisingly, approximately half of the A2/MART127-35 TCRs derived from CD4(+) T cells, but none from CD8(+) T cells, were stained by A2/MART127-35 monomer and possessed broader cross-reactivity. Our results suggest that the differences in the primary structure of peripheral CD4(+) and CD8(+) TCRβ repertoire indeed result in the differences in their ability to form extraordinarily high avidity T cells which would otherwise have been deleted by central tolerance.

CD4(+) and CD8(+) TCRβ repertoires possess different potentials to generate extraordinarily high-avidity T cells.

Invariant natural killer T (iNKT) cells recognize self-lipids presented by CD1d through characteristic TCRs, which mainly consist of the invariant Vα14-Jα18 TCRα chain and Vβ8.2, 7 or 2 TCRβ chains with hypervariable CDR3β sequences in mice. The iNKT cell-CD1d axis is conserved between humans and mice, and human CD1d reactivity of murine iNKT cells have been described. However, the detailed differences between the recognition of human and mouse CD1d bound to various self-lipids by mouse iNKT TCRs are largely unknown. In this study, we generated a de novo murine iNKT TCR repertoire with a wider range of autoreactivity compared with that of naturally occurring peripheral iNKT TCRs. Vβ8.2 mouse iNKT TCRs capable of recognizing the human CD1d-self-lipid tetramer were identified, although such clones were not detectable in the Vβ7 or Vβ2 iNKT TCR repertoire. In line with previously reports, clonotypic Vβ8.2 iNKT TCRs with unique CDR3β loops did not discriminate among lipids presented by mouse CD1d. Unexpectedly, however, these iNKT TCRs showed greater ligand selectivity toward human CD1d presenting the same lipids. Our findings demonstrated that the recognition of mouse and human CD1d-self-lipid complexes by murine iNKT TCRs is not conserved, thereby further elucidating the differences between cognate and cross-species reactivity of self-antigens by mouse iNKT TCRs.

Mouse and Human CD1d-Self-Lipid Complexes Are Recognized Differently by Murine Invariant Natural Killer T Cell Receptors.

Adoptive immunotherapy is a potentially curative therapeutic approach for patients with advanced cancer. However, the in vitro expansion of antitumor T cells prior to infusion inevitably incurs differentiation towards effector T cells and impairs persistence following adoptive transfer. Epigenetic profiles regulate gene expression of key transcription factors over the course of immune cell differentiation, proliferation, and function. Using comprehensive screening of chemical probes with defined epigenetic targets, we found that JQ1, an inhibitor of bromodomain and extra-terminal motif (BET) proteins, maintained CD8+ T cells with functional properties of stem cell-like and central memory T cells. Mechanistically, the BET protein BRD4 directly regulated expression of the transcription factor BATF in CD8+ T cells, which was associated with differentiation of T cells into an effector memory phenotype. JQ1-treated T cells showed enhanced persistence and antitumor effects in murine T cell receptor and chimeric antigen receptor gene therapy models. Furthermore, we found that histone acetyltransferase p300 supported the recruitment of BRD4 to the BATF promoter region, and p300 inhibition similarly augmented antitumor effects of the adoptively transferred T cells. These results demonstrate that targeting the BRD4-p300 signaling cascade supports the generation of superior antitumor T cell grafts for adoptive immunotherapy.

Article	Year
CD4(+) and CD8(+) TCRβ repertoires possess different potentials to generate extraordinarily high-avidity T cells. Scientific reports\| PubMed ID: 27030642	2016
Mouse and Human CD1d-Self-Lipid Complexes Are Recognized Differently by Murine Invariant Natural Killer T Cell Receptors. PloS one\| PubMed ID: 27213277	2016
BET bromodomain inhibition enhances T cell persistence and function in adoptive immunotherapy models. The Journal of clinical investigation\| PubMed ID: 27548527	2016

Mark Anczurowski

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