Redirecting T cells to ewing's sarcoma family of tumors by a chimeric NKG2D receptor expressed by lentiviral transduction or mRNA transfection

Lehner M, Goetz G, Proff J, Schaft N, Dörrie J, Full F, Enßer A, Muller Y, Cerwenka A, Abken H, Parolini O, Ambros PF, Kovar H, Holter W (2012)

Publication Type: Journal article

Publication year: 2012

Journal

Book Volume: 7

Article Number: e31210

Journal Issue: 2

DOI: 10.1371/journal.pone.0031210

Abstract

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. © 2012 Lehner et al.

Authors with CRIS profile

Involved external institutions

Excellence of Fondazione Poliambulanza Italy (IT) Deutsches Krebsforschungszentrum (DKFZ) Germany (DE) Universität zu Köln Germany (DE) St. Anna Kinderspital Austria (AT)

How to cite

APA:

Lehner, M., Goetz, G., Proff, J., Schaft, N., Dörrie, J., Full, F.,... Holter, W. (2012). Redirecting T cells to ewing's sarcoma family of tumors by a chimeric NKG2D receptor expressed by lentiviral transduction or mRNA transfection. PLoS ONE, 7(2). https://dx.doi.org/10.1371/journal.pone.0031210

MLA:

Lehner, Manfred, et al. "Redirecting T cells to ewing's sarcoma family of tumors by a chimeric NKG2D receptor expressed by lentiviral transduction or mRNA transfection." PLoS ONE 7.2 (2012).

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