Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Identification of human T-cell receptors with optimal affinity to cancer antigens using antigen-negative humanized mice

Nature Biotechnology volume 33, pages 402–407 (2015)Cite this article

Subjects

Abstract

Identifying T-cell receptors (TCRs) that bind tumor-associated antigens (TAAs) with optimal affinity is a key bottleneck in the development of adoptive T-cell therapy of cancer1. TAAs are unmutated self proteins, and T cells bearing high-affinity TCRs specific for such antigens are commonly deleted in the thymus2. To identify optimal-affinity TCRs, we generated antigen-negative humanized mice with a diverse human TCR repertoire restricted to the human leukocyte antigen (HLA) A*02:01 (ref. 3). These mice were immunized with human TAAs, for which they are not tolerant, allowing induction of CD8+ T cells with optimal-affinity TCRs. We isolate TCRs specific for the cancer/testis (CT) antigen MAGE-A1 (ref. 4) and show that two of them have an anti-tumor effect in vivo. By comparison, human-derived TCRs have lower affinity and do not mediate substantial therapeutic effects. We also identify optimal-affinity TCRs specific for the CT antigen NY-ESO. Our humanized mouse model provides a useful tool for the generation of optimal-affinity TCRs for T-cell therapy.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Learn more

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Generation of MAGE-A1-specific TCRs in ABabDII mice.
Figure 2: Specificity of ABabDII derived TCRs.
Figure 3: Functional comparison of ABabDII-derived TCRs with a human-derived TCR in vitro and in vivo.
Figure 4: Functional comparison of a NY-ESO157-specific TCR from ABabDII mice with a patient-derived TCR.

Accession codes

Primary accessions

NCBI Reference Sequence

References

  1. June, C.H. Adoptive T cell therapy for cancer in the clinic. J. Clin. Invest. 117, 1466–1476 (2007).

    Article  CAS  Google Scholar 

  2. Theobald, M. et al. Tolerance to p53 by A2.1-restricted cytotoxic T lymphocytes. J. Exp. Med. 185, 833–841 (1997).

    Article  CAS  Google Scholar 

  3. Li, L.-P. et al. Transgenic mice with a diverse human T cell antigen receptor repertoire. Nat. Med. 16, 1029–1034 (2010).

    Article  CAS  Google Scholar 

  4. van der Bruggen, P. et al. A gene encoding an antigen recognized by cytolytic T lymphocytes on a human melanoma. Science 254, 1643–1647 (1991).

    Article  CAS  Google Scholar 

  5. Holler, P.D., Chlewicki, L.K. & Kranz, D.M. TCRs with high affinity for foreign pMHC show self-reactivity. Nat. Immunol. 4, 55–62 (2003).

    Article  CAS  Google Scholar 

  6. Linette, G.P. et al. Cardiovascular toxicity and titin cross-reactivity of affinity-enhanced T cells in myeloma and melanoma. Blood 122, 863–871 (2013).

    Article  CAS  Google Scholar 

  7. Engels, B., Chervin, A.S., Sant, A.J., Kranz, D.M. & Schreiber, H. Long-term persistence of CD4(+) but rapid disappearance of CD8(+) T cells expressing an MHC class I-restricted TCR of nanomolar affinity. Mol. Ther. 20, 652–660 (2012).

    Article  CAS  Google Scholar 

  8. Sadovnikova, E. & Stauss, H.J. Peptide-specific cytotoxic T lymphocytes restricted by nonself major histocompatibility complex class I molecules: reagents for tumor immunotherapy. Proc. Natl. Acad. Sci. USA 93, 13114–13118 (1996).

    Article  CAS  Google Scholar 

  9. Johnson, L.A. et al. Gene therapy with human and mouse T-cell receptors mediates cancer regression and targets normal tissues expressing cognate antigen. Blood 114, 535–546 (2009).

    Article  CAS  Google Scholar 

  10. Parkhurst, M.R. et al. Characterization of genetically modified T-cell receptors that recognize the CEA:691–699 peptide in the context of HLA-A2.1 on human colorectal cancer cells. Clin. Cancer Res. 15, 169–180 (2009).

    Article  CAS  Google Scholar 

  11. Chinnasamy, N. et al. A TCR targeting the HLA-A*0201-restricted epitope of MAGE-A3 recognizes multiple epitopes of the MAGE-A antigen superfamily in several types of cancer. J. Immunol. 186, 685–696 (2011).

    Article  CAS  Google Scholar 

  12. Davis, J.L. et al. Development of human anti-murine T-cell receptor antibodies in both responding and nonresponding patients enrolled in TCR gene therapy trials. Clin. Cancer Res. 16, 5852–5861 (2010).

    Article  CAS  Google Scholar 

  13. Offringa, R. Antigen choice in adoptive T-cell therapy of cancer. Curr. Opin. Immunol. 21, 190–199 (2009).

    Article  CAS  Google Scholar 

  14. Pascolo, S. et al. A MAGE-A1 HLA-A A*0201 epitope identified by mass spectrometry. Cancer Res. 61, 4072–4077 (2001).

    CAS  PubMed  Google Scholar 

  15. Toso, J.F. et al. MAGE-1-specific precursor cytotoxic T-lymphocytes present among tumor-infiltrating lymphocytes from a patient with breast cancer: characterization and antigen-specific activation. Cancer Res. 56, 16–20 (1996).

    CAS  PubMed  Google Scholar 

  16. Ottaviani, S., Zhang, Y., Boon, T. & van der Bruggen, P.A. MAGE-1 antigenic peptide recognized by human cytolytic T lymphocytes on HLA-A2 tumor cells. Cancer Immunol. Immunother. 54, 1214–1220 (2005).

    Article  CAS  Google Scholar 

  17. Huijbers, I.J. et al. Minimal tolerance to a tumor antigen encoded by a cancer-germline gene. J. Immunol. 188, 111–121 (2012).

    Article  CAS  Google Scholar 

  18. Lee, E.-C. et al. Complete humanization of the mouse immunoglobulin loci enables efficient therapeutic antibody discovery. Nat. Biotechnol. 32, 356–363 (2014).

    Article  CAS  Google Scholar 

  19. Sommermeyer, D. & Uckert, W. Minimal amino acid exchange in human TCR constant regions fosters improved function of TCR gene-modified T cells. J. Immunol. 184, 6223–6231 (2010).

    Article  CAS  Google Scholar 

  20. Morgan, R.A. et al. Cancer regression and neurological toxicity following anti-MAGE-A3 TCR gene therapy. J. Immunother. 36, 133–151 (2013).

    Article  CAS  Google Scholar 

  21. Linnemann, C. et al. High-throughput identification of antigen-specific TCRs by TCR gene capture. Nat. Med. 19, 1534–1541 (2013).

    Article  CAS  Google Scholar 

  22. Chen, J.-L. et al. Structural and kinetic basis for heightened immunogenicity of T cell vaccines. J. Exp. Med. 201, 1243–1255 (2005).

    Article  CAS  Google Scholar 

  23. Vierboom, M.P. et al. Tumor eradication by wild-type p53-specific cytotoxic T lymphocytes. J. Exp. Med. 186, 695–704 (1997).

    Article  CAS  Google Scholar 

  24. Scott-Browne, J.P., White, J., Kappler, J.W., Gapin, L. & Marrack, P. Germline-encoded amino acids in the alphabeta T-cell receptor control thymic selection. Nature 458, 1043–1046 (2009).

    Article  CAS  Google Scholar 

  25. Aleksic, M. et al. Different affinity windows for virus and cancer-specific T-cell receptors: implications for therapeutic strategies. Eur. J. Immunol. 42, 3174–3179 (2012).

    Article  CAS  Google Scholar 

  26. Schmid, D.A. et al. Evidence for a TCR affinity threshold delimiting maximal CD8 T cell function. J. Immunol. 184, 4936–4946 (2010).

    Article  CAS  Google Scholar 

  27. Wang, B. et al. A single peptide-MHC complex positively selects a diverse and specific CD8 T cell repertoire. Science 326, 871–874 (2009).

    Article  CAS  Google Scholar 

  28. Birnbaum, M.E. et al. Deconstructing the peptide-MHC specificity of T cell recognition. Cell 157, 1073–1087 (2014).

    Article  CAS  Google Scholar 

  29. Gotter, J., Brors, B., Hergenhahn, M. & Kyewski, B. Medullary epithelial cells of the human thymus express a highly diverse selection of tissue-specific genes colocalized in chromosomal clusters. J. Exp. Med. 199, 155–166 (2004).

    Article  CAS  Google Scholar 

  30. Kammertoens, T. & Blankenstein, T. It's the peptide-MHC affinity, stupid. Cancer Cell 23, 429–431 (2013).

    Article  CAS  Google Scholar 

  31. Huang, L.Q. et al. Cytolytic T lymphocytes recognize an antigen encoded by MAGE-A10 on a human melanoma. J. Immunol. 162, 6849–6854 (1999).

    CAS  PubMed  Google Scholar 

  32. Zarour, H. et al. The majority of autologous cytolytic T-lymphocyte clones derived from peripheral blood lymphocytes of a melanoma patient recognize an antigenic peptide derived from Gene Pmel17/gp100. J. Invest. Dermatol. 107, 63–67 (1996).

    Article  CAS  Google Scholar 

  33. Hérin, M. et al. Production of stable cytolytic T-cell clones directed against autologous human melanoma. Int. J. Cancer 39, 390–396 (1987).

    Article  Google Scholar 

  34. Weynants, P. et al. Derivation of tumor-specific cytolytic T-cell clones from two lung cancer patients with long survival. Am. J. Respir. Crit. Care Med. 159, 55–62 (1999).

    Article  CAS  Google Scholar 

  35. Pellat-Deceunynck, C. et al. The cancer germ-line genes MAGE-1, MAGE-3 and PRAME are commonly expressed by human myeloma cells. Eur. J. Immunol. 30, 803–809 (2000).

    Article  CAS  Google Scholar 

  36. Bredenbeck, A. et al. Coordinated expression of clustered cancer/testis genes encoded in a large inverted repeat DNA structure. Gene 415, 68–73 (2008).

    Article  CAS  Google Scholar 

  37. Sommermeyer, D. et al. NY-ESO-1 antigen-reactive T cell receptors exhibit diverse therapeutic capability. Int. J. Cancer 132, 1360–1367 (2013).

    Article  CAS  Google Scholar 

  38. Morita, S., Kojima, T. & Kitamura, T. Plat-E: an efficient and stable system for transient packaging of retroviruses. Gene Ther. 7, 1063–1066 (2000).

    Article  CAS  Google Scholar 

  39. Leisegang, M. et al. Enhanced functionality of T cell receptor-redirected T cells is defined by the transgene cassette. J. Mol. Med. (Berl) 86, 573–583 (2008).

    Article  CAS  Google Scholar 

  40. Kuball, J. et al. Facilitating matched pairing and expression of TCR chains introduced into human T cells. Blood 109, 2331–2338 (2007).

    Article  CAS  Google Scholar 

  41. Cohen, C.J. et al. Enhanced antitumor activity of T cells engineered to express T-cell receptors with a second disulfide bond. Cancer Res. 67, 3898–3903 (2007).

    Article  CAS  Google Scholar 

  42. Engels, B. et al. Retroviral vectors for high-level transgene expression in T lymphocytes. Hum. Gene Ther. 14, 1155–1168 (2003).

    Article  CAS  Google Scholar 

  43. Uckert, W. et al. Efficient gene transfer into primary human CD8+ T lymphocytes by MuLV-10A1 retrovirus pseudotype. Hum. Gene Ther. 11, 1005–1014 (2000).

    Article  CAS  Google Scholar 

  44. Heemskerk, M.H.M. et al. Redirection of antileukemic reactivity of peripheral T lymphocytes using gene transfer of minor histocompatibility antigen HA-2-specific T-cell receptor complexes expressing a conserved alpha joining region. Blood 102, 3530–3540 (2003).

    Article  CAS  Google Scholar 

  45. Wilde, S. et al. Dendritic cells pulsed with RNA encoding allogeneic MHC and antigen induce T cells with superior antitumor activity and higher TCR functional avidity. Blood 114, 2131–2139 (2009).

    Article  CAS  Google Scholar 

  46. de Castro, E. et al. ScanProsite: detection of PROSITE signature matches and ProRule-associated functional and structural residues in proteins. Nucleic Acids Res. 34, W362–W365 (2006).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

The authors thank G. Willimsky for discussion, S. Kupsch and S. Fürl for technical assistance, and I. Hoeft for animal caretaking. C. Linnemann and T. Schumacher kindly provided the NY-ESO-1157/HLA-A2 multimer. This work was supported by the Deutsche Forschungsgemeinschaft (Sonderforschungsbereich TR36) and the Berlin Institute of Health (BIH).

Author information

Author notes

  1. Catarina Leitão

    Present address: Present address: Institute for Molecular and Cell Biology, Porto, Portugal.,

Authors and Affiliations

  1. Max-Delbrück-Center for Molecular Medicine, Berlin, Germany

    Matthias Obenaus, Catarina Leitão, Matthias Leisegang, Xiaojing Chen, Ioannis Gavvovidis, Wolfgang Uckert & Thomas Blankenstein

  2. Ludwig Institute for Cancer Research and WELBIO, Brussels, Belgium

    Pierre van der Bruggen

  3. De Duve Institute, Université Catholique de Louvain, Brussels, Belgium

    Pierre van der Bruggen

  4. Institute of Biology, Humboldt University, Berlin, Germany

    Wolfgang Uckert

  5. Medigene AG, Planegg/Martinsried, Germany

    Dolores J Schendel

  6. Institute of Immunology, Charité Campus Buch, Berlin, Germany

    Thomas Blankenstein

Authors
  1. Matthias Obenaus
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Catarina Leitão
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Matthias Leisegang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Xiaojing Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Ioannis Gavvovidis
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Pierre van der Bruggen
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Wolfgang Uckert
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Dolores J Schendel
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Thomas Blankenstein
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

M.O. planned and performed most experiments, analyzed data and wrote the manuscript. C.L. initiated the project, established new methods, planned and performed experiments, and analyzed data. M.L., X.C. and I.G. planned and performed experiments, and analyzed data. P.v.d.B. and D.J.S. provided unique reagents and advised on their use. W.U. planned experiments and analyzed data. T.B. supervised the project, analyzed data and wrote the manuscript. All authors revised the manuscript.

Corresponding author

Correspondence to Thomas Blankenstein.

Ethics declarations

Competing interests

The Max-Delbrück-Center for Molecular Medicine (T.B., M.O., C.L.) applied for a patent on the MAGE-A1-specific TCRs.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–8 and Supplementary Tables 1–3 (PDF 1049 kb)

Rights and permissions

Reprints and Permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Obenaus, M., Leitão, C., Leisegang, M. et al. Identification of human T-cell receptors with optimal affinity to cancer antigens using antigen-negative humanized mice. Nat Biotechnol 33, 402–407 (2015). https://doi.org/10.1038/nbt.3147

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nbt.3147

This article is cited by

Search

Advanced search

Quick links

Nature Briefing: Translational Research

Sign up for the Nature Briefing: Translational Research newsletter — top stories in biotechnology, drug discovery and pharma.

Get what matters in translational research, free to your inbox weekly. Sign up for Nature Briefing: Translational Research