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Redirecting T Cells to Ewing's Sarcoma Family of Tumors by a Chimeric NKG2D Receptor Expressed by Lentiviral Transduction or mRNA Transfection

We explored the possibility to target Ewing's sarcoma family of tumors (ESFT) by redirecting T cells. To this aim, we considered NKG2D-ligands (NKG2D-Ls) as possible target antigens. Detailed analysis of the expression of MICA, MICB, ULBP-1, -2, and -3 in fourteen ESFT cell lines revealed consi...

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Autores principales: Lehner, Manfred, Götz, Gabriel, Proff, Julia, Schaft, Niels, Dörrie, Jan, Full, Florian, Ensser, Armin, Muller, Yves A., Cerwenka, Adelheid, Abken, Hinrich, Parolini, Ornella, Ambros, Peter F., Kovar, Heinrich, Holter, Wolfgang
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3280271/
https://www.ncbi.nlm.nih.gov/pubmed/22355347
http://dx.doi.org/10.1371/journal.pone.0031210
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author Lehner, Manfred
Götz, Gabriel
Proff, Julia
Schaft, Niels
Dörrie, Jan
Full, Florian
Ensser, Armin
Muller, Yves A.
Cerwenka, Adelheid
Abken, Hinrich
Parolini, Ornella
Ambros, Peter F.
Kovar, Heinrich
Holter, Wolfgang
author_facet Lehner, Manfred
Götz, Gabriel
Proff, Julia
Schaft, Niels
Dörrie, Jan
Full, Florian
Ensser, Armin
Muller, Yves A.
Cerwenka, Adelheid
Abken, Hinrich
Parolini, Ornella
Ambros, Peter F.
Kovar, Heinrich
Holter, Wolfgang
author_sort Lehner, Manfred
collection PubMed
description We explored the possibility to target Ewing's sarcoma family of tumors (ESFT) by redirecting T cells. To this aim, we considered NKG2D-ligands (NKG2D-Ls) as possible target antigens. Detailed analysis of the expression of MICA, MICB, ULBP-1, -2, and -3 in fourteen ESFT cell lines revealed consistent expression of at least one NKG2D-L. Thus, for redirecting T cells, we fused a CD3ζ/CD28-derived signaling domain to the ectodomain of NKG2D, however, opposite transmembrane orientation of this signaling domain and NKG2D required inverse orientation fusion of either of them. We hypothesized that the particularly located C-terminus of the NKG2D ectodomain should allow reengineering of the membrane anchoring from a native N-terminal to an artificial C-terminal linkage. Indeed, the resulting chimeric NKG2D receptor (chNKG2D) was functional and efficiently mediated ESFT cell death triggered by activated T cells. Notably, ESFT cells with even low NKG2D-L expression were killed by CD8(pos) and also CD4(pos) cells. Both, mRNA transfection and lentiviral transduction resulted in high level surface expression of chNKG2D. However, upon target-cell recognition receptor surface levels were maintained by tranfected RNA only during the first couple of hours after transfection. Later, target-cell contact resulted in strong and irreversible receptor down-modulation, whereas lentivirally mediated expression of chNKG2D remained constant under these conditions. Together, our study defines NKG2D-Ls as targets for a CAR-mediated T cell based immunotherapy of ESFT. A comparison of two different methods of gene transfer reveals strong differences in the susceptibility to ligand-induced receptor down-modulation with possible implications for the applicability of RNA transfection.
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spelling pubmed-32802712012-02-21 Redirecting T Cells to Ewing's Sarcoma Family of Tumors by a Chimeric NKG2D Receptor Expressed by Lentiviral Transduction or mRNA Transfection Lehner, Manfred Götz, Gabriel Proff, Julia Schaft, Niels Dörrie, Jan Full, Florian Ensser, Armin Muller, Yves A. Cerwenka, Adelheid Abken, Hinrich Parolini, Ornella Ambros, Peter F. Kovar, Heinrich Holter, Wolfgang PLoS One Research Article We explored the possibility to target Ewing's sarcoma family of tumors (ESFT) by redirecting T cells. To this aim, we considered NKG2D-ligands (NKG2D-Ls) as possible target antigens. Detailed analysis of the expression of MICA, MICB, ULBP-1, -2, and -3 in fourteen ESFT cell lines revealed consistent expression of at least one NKG2D-L. Thus, for redirecting T cells, we fused a CD3ζ/CD28-derived signaling domain to the ectodomain of NKG2D, however, opposite transmembrane orientation of this signaling domain and NKG2D required inverse orientation fusion of either of them. We hypothesized that the particularly located C-terminus of the NKG2D ectodomain should allow reengineering of the membrane anchoring from a native N-terminal to an artificial C-terminal linkage. Indeed, the resulting chimeric NKG2D receptor (chNKG2D) was functional and efficiently mediated ESFT cell death triggered by activated T cells. Notably, ESFT cells with even low NKG2D-L expression were killed by CD8(pos) and also CD4(pos) cells. Both, mRNA transfection and lentiviral transduction resulted in high level surface expression of chNKG2D. However, upon target-cell recognition receptor surface levels were maintained by tranfected RNA only during the first couple of hours after transfection. Later, target-cell contact resulted in strong and irreversible receptor down-modulation, whereas lentivirally mediated expression of chNKG2D remained constant under these conditions. Together, our study defines NKG2D-Ls as targets for a CAR-mediated T cell based immunotherapy of ESFT. A comparison of two different methods of gene transfer reveals strong differences in the susceptibility to ligand-induced receptor down-modulation with possible implications for the applicability of RNA transfection. Public Library of Science 2012-02-15 /pmc/articles/PMC3280271/ /pubmed/22355347 http://dx.doi.org/10.1371/journal.pone.0031210 Text en Lehner et al. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Lehner, Manfred
Götz, Gabriel
Proff, Julia
Schaft, Niels
Dörrie, Jan
Full, Florian
Ensser, Armin
Muller, Yves A.
Cerwenka, Adelheid
Abken, Hinrich
Parolini, Ornella
Ambros, Peter F.
Kovar, Heinrich
Holter, Wolfgang
Redirecting T Cells to Ewing's Sarcoma Family of Tumors by a Chimeric NKG2D Receptor Expressed by Lentiviral Transduction or mRNA Transfection
title Redirecting T Cells to Ewing's Sarcoma Family of Tumors by a Chimeric NKG2D Receptor Expressed by Lentiviral Transduction or mRNA Transfection
title_full Redirecting T Cells to Ewing's Sarcoma Family of Tumors by a Chimeric NKG2D Receptor Expressed by Lentiviral Transduction or mRNA Transfection
title_fullStr Redirecting T Cells to Ewing's Sarcoma Family of Tumors by a Chimeric NKG2D Receptor Expressed by Lentiviral Transduction or mRNA Transfection
title_full_unstemmed Redirecting T Cells to Ewing's Sarcoma Family of Tumors by a Chimeric NKG2D Receptor Expressed by Lentiviral Transduction or mRNA Transfection
title_short Redirecting T Cells to Ewing's Sarcoma Family of Tumors by a Chimeric NKG2D Receptor Expressed by Lentiviral Transduction or mRNA Transfection
title_sort redirecting t cells to ewing's sarcoma family of tumors by a chimeric nkg2d receptor expressed by lentiviral transduction or mrna transfection
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3280271/
https://www.ncbi.nlm.nih.gov/pubmed/22355347
http://dx.doi.org/10.1371/journal.pone.0031210
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