Effective screening of T cells recognizing neoantigens and construction of T-cell receptor-engineered T cells
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Taigo Kato1,*, Tatsuo Matsuda1,*, Yuji Ikeda1,*, Jae-Hyun Park1, Matthias Leisegang2,3, Sachiko Yoshimura4, Tetsuro Hikichi4, Makiko Harada4, Makda Zewde1, Sho Sato5, Kosei Hasegawa5, Kazuma Kiyotani1 and Yusuke Nakamura1,6
1Department of Medicine, The University of Chicago, Chicago, IL 60637, USA
2Institute of Immunology – Campus Buch, Charité – Universitätsmedizin Berlin, Berlin 13125, Germany
3Berlin Institute of Health, Berlin 10117, Germany
4OncoTherapy Science Inc., Kawasaki, Kanagawa 213-0012, Japan
5Department of Gynecologic Oncology, Saitama Medical University International Medical Center, Hidaka, Saitama 350-1298, Japan
6Department of Surgery, The University of Chicago, Chicago, IL 60637, USA
*These authors contributed equally to this work
Yusuke Nakamura, email: [email protected]
Keywords: neoantigens, neoantigen-specific T cells, T cell receptor, TCR-engineered T cells
Received: November 17, 2017 Accepted: January 09, 2018 Published: January 13, 2018
Neoantigens are the main targets of tumor-specific T cells reactivated by immune checkpoint-blocking antibodies or when using tumor-infiltrating T cells for adoptive therapy. While cancers often accumulate hundreds of mutations and harbor several immunogenic neoantigens, the repertoire of mutation-specific T cells in patients might be restricted. To bypass suboptimal conditions, which impede the reactivation of existing T cells or the priming of neoantigen-specific T cells in a patient, we employ T cells of healthy donors with an overlapping HLA repertoire to target cancer neoantigens. In this study, we focus on streamlining the process of in vitro-induction of neoantigen-specific T cells and isolating their T cell receptors (TCRs) to establish a time-efficient protocol that will allow the patient to benefit from subsequent therapy. We first optimized the priming of T cells to omit multiple restimulations and extended culturing. Neoantigen-specific T cells were enriched using specific dextramers and next-generation sequencing was applied to determine the TCR repertoire. This allowed us to circumvent the laborious process of expanding T cell clones. Using this protocol, we successfully identified HLA-A-restricted TCRs specific for neoantigens found in an esophageal cancer cell line (TE-8) and a primary ovarian cancer. To verify TCR specificity, we generated TCR-engineered T cells and confirmed recognition of the tumor-derived neoantigens. Our results also emphasize the importance of neoepitope selection in order to avoid cross-reactivity to corresponding wild-type peptide sequences. In conclusion, we established a 2-week protocol for generating and identifying neoantigen-specific TCRs from third-party donors making this strategy applicable for clinical use.
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