An optimized single chain TCR scaffold relying on the assembly with the native CD3-complex prevents residual mispairing with endogenous TCRs in human T-cells.

Details

Ressource 1Download: BIB_45D1E4A5393B.P001.pdf (4939.20 [Ko])
State: Public
Version: author
Serval ID
serval:BIB_45D1E4A5393B
Type
Article: article from journal or magazin.
Collection
Publications
Institution
Title
An optimized single chain TCR scaffold relying on the assembly with the native CD3-complex prevents residual mispairing with endogenous TCRs in human T-cells.
Journal
Oncotarget
Author(s)
Knies D., Klobuch S., Xue S.A., Birtel M., Echchannaoui H., Yildiz O., Omokoko T., Guillaume P., Romero P., Stauss H., Sahin U., Herr W., Theobald M., Thomas S., Voss R.H.
ISSN
1949-2553 (Electronic)
ISSN-L
1949-2553
Publication state
Published
Issued date
19/04/2016
Peer-reviewed
Oui
Volume
7
Number
16
Pages
21199-21221
Language
english
Notes
Publication types: Journal Article
Publication Status: ppublish
Abstract
Immunotherapy of cancer envisions the adoptive transfer of T-cells genetically engineered with tumor-specific heterodimeric α/β T-cell receptors (TCRα/β). However, potential mispairing of introduced TCRα/β-chains with endogenous β/α-ones may evoke unpredictable autoimmune reactivities. A novel single chain (sc)TCR format relies on the fusion of the Vα-Linker-Vβ-fragment to the TCR Cβ-domain and coexpression of the TCR Cα-domain capable of recruiting the natural CD3-complex for full and hence, native T-cell signaling. Here, we tested whether such a gp100(280-288)- or p53(264-272) tumor antigen-specific scTCR is still prone to mispairing with TCRα. In a human Jurkat-76 T-cell line lacking endogenous TCRs, surface expression and function of a scTCR could be reconstituted by any cointroduced TCRα-chain indicating mispairing to take place on a molecular basis. In contrast, transduction into human TCRα/β-positive T-cells revealed that mispairing is largely reduced. Competition experiments in Jurkat-76 confirmed the preference of dcTCR to selfpair and to spare scTCR. This also allowed for the generation of dc/scTCR-modified cytomegalovirus/tumor antigen-bispecific T-cells to augment T-cell activation in CMV-infected tumor patients. Residual mispairing was prevented by strenghtening the Vα-Li-Vβ-fragment through the design of a novel disulfide bond between a Vα- and a linker-resident residue close to Vβ. Multimer-stainings, and cytotoxicity-, IFNγ-secretion-, and CFSE-proliferation-assays, the latter towards dendritic cells endogenously processing RNA-electroporated gp100 antigen proved the absence of hybrid scTCR/TCRα-formation without impairing avidity of scTCR/Cα in T-cells. Moreover, a fragile cytomegalovirus pp65(495-503)-specific scTCR modified this way acquired enhanced cytotoxicity. Thus, optimized scTCR/Cα inhibits residual TCR mispairing to accomplish safe adoptive immunotherapy for bulk endogenous TCRα/β-positive T-cells.

Keywords
Adoptive Transfer, Animals, Biomarkers, Tumor, CD3 Complex/genetics, CD3 Complex/immunology, Cell Membrane, Cell Proliferation, Humans, Immunotherapy, Adoptive, Leukemia, T-Cell/genetics, Leukemia, T-Cell/immunology, Leukemia, T-Cell/pathology, Mice, Receptors, Antigen, T-Cell, alpha-beta/genetics, Receptors, Antigen, T-Cell, alpha-beta/immunology, Signal Transduction, T-Lymphocytes/immunology, T-Lymphocytes/metabolism, Tumor Cells, Cultured, Immune response, Immunity, Immunology and Microbiology Section, T-cell receptors, T-cells, gene therapy, human, tumor immunity
Pubmed
Web of science
Open Access
Yes
Create date
09/04/2016 15:02
Last modification date
20/08/2019 13:50
Usage data