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Retargeting NK92 cells using an HLA-A2-restricted, EBNA3C-specific chimeric antigen receptor

Abstract

Advances in adoptive cell immunotherapy have led to several promising options for cancer patients. Single-chain variable fragments (scFvs) were isolated from a human phage display library by panning on recombinant human leukocyte antigen (HLA)-A2-peptide complexes. A scFv (EBNA Clone 315) specific for HLA-A2 carrying a 10 amino acid peptide (LLDFVRFMGV) derived from the Epstein–Barr virus latent protein EBNA3C was fully characterized. EBNA Clone 315 displayed exquisite specificity toward its targeted T-cell epitope (TCE) and did not cross-react with the free peptide, HLA-A2 complexes, which carried irrelevant peptides, or HLA-A2 cells. Furthermore, after engineering into a scFv–Fc fusion protein, we were able to determine its affinity, detection sensitivity, and ability to induce antibody-dependent cellular cytotoxicity (ADCC). As a proof-of-principle, a chimeric antigen receptor (CAR) version of EBNA Clone 315 was used to reprogram NK92MI cells. CAR-expressing NK92MI cells showed highly specific and potent cytotoxicity toward the targeted TCE, with detection sensitivity of approximately 25 molecules and cytolytic capacity threefold greater than scFv-Fc-mediated ADCC. For the first time, we show the successful reprogramming of non-T cells toward a specific TCE using a CAR.

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Acknowledgements

We would like to thank the team of Eric Pamer, along with Ingrid Leiner, at the MSKCC Tetramer facility for preparing all the HLA-A2-peptide complexes used in this study. We would also like to thank Richard O’Reilly and Lorna Barnett for providing us with the BLCL lines. We would also like to thank Hong Xu and Hong-fen Guo in our laboratory for help with constructing the scFv–Fc expression vector. This work was partly supported by the Catie Hoch Fund, Katies Find-A-Cure Fund, Robert Steele Laboratory Fund and the Caitlin Robb Foundation.

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Tassev, D., Cheng, M. & Cheung, NK. Retargeting NK92 cells using an HLA-A2-restricted, EBNA3C-specific chimeric antigen receptor. Cancer Gene Ther 19, 84–100 (2012). https://doi.org/10.1038/cgt.2011.66

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Keywords

  • HLA-A2
  • NK92MI
  • ScFv
  • ScFv-Fc
  • chimeric antigen receptors
  • antibodies

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