Skip to main content

Thank you for visiting 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.

Mining exomic sequencing data to identify mutated antigens recognized by adoptively transferred tumor-reactive T cells

This article has been updated


Substantial regressions of metastatic lesions have been observed in up to 70% of patients with melanoma who received adoptively transferred autologous tumor-infiltrating lymphocytes (TILs) in phase 2 clinical trials1,2. In addition, 40% of patients treated in a recent trial experienced complete regressions of all measurable lesions for at least 5 years following TIL treatment3. To evaluate the potential association between the ability of TILs to mediate durable regressions and their ability to recognize potent antigens that presumably include mutated gene products, we developed a new screening approach involving mining whole-exome sequence data to identify mutated proteins expressed in patient tumors. We then synthesized and evaluated candidate mutated T cell epitopes that were identified using a major histocompatibility complex–binding algorithm4 for recognition by TILs. Using this approach, we identified mutated antigens expressed on autologous tumor cells that were recognized by three bulk TIL lines from three individuals with melanoma that were associated with objective tumor regressions following adoptive transfer. This simplified approach for identifying mutated antigens recognized by T cells avoids the need to generate and laboriously screen cDNA libraries from tumors and may represent a generally applicable method for identifying mutated antigens expressed in a variety of tumor types.

This is a preview of subscription content

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: Response of TIL 2098 to candidate epitopes identified from autologous tumors.
Figure 2: Response of TIL 2369 to candidate epitopes identified from autologous tumors.
Figure 3: Response of TIL 3309 to candidate epitopes identified from autologous tumors.
Figure 4: IFN-γ ELISPOT responses of TIL and PBMCs obtained before and after autologous TIL transfer.

Change history

  • 14 May 2013

     In the version of this article initially published online, the second sentence of the abstract stated, “In addition, 40% of patients treated in a recent trial experienced complete regressions of all measurable lesions lasting between 5 and 9 years after treatment3.” The correct statement should read, “In addition, 40% of patients treated in a recent trial experienced complete regressions of all measurable lesions for at least 5 years following TIL treatment3.” The error has been corrected for the print, PDF and HTML versions of this article.


  1. 1

    Dudley, M.E. et al. Cancer regression and autoimmunity in patients after clonal repopulation with antitumor lymphocytes. Science 298, 850–854 (2002).

    CAS  Article  Google Scholar 

  2. 2

    Dudley, M.E. et al. Adoptive cell therapy for patients with metastatic melanoma: evaluation of intensive myeloablative chemoradiation preparative regimens. J. Clin. Oncol. 26, 5233–5239 (2008).

    CAS  Article  Google Scholar 

  3. 3

    Rosenberg, S.A. et al. Durable complete responses in heavily pretreated patients with metastatic melanoma using T-cell transfer immunotherapy. Clin. Cancer Res. 17, 4550–4557 (2011).

    CAS  Article  Google Scholar 

  4. 4

    Nielsen, M. et al. NetMHCpan, a method for quantitative predictions of peptide binding to any HLA-A and -B locus protein of known sequence. PLoS ONE 2, e796 (2007).

    Article  Google Scholar 

  5. 5

    Marincola, F.M. et al. Locus-specific analysis of human leukocyte antigen class I expression in melanoma cell lines. J. Immunother. Emphasis Tumor Immunol. 16, 13–23 (1994).

    CAS  Article  Google Scholar 

  6. 6

    Salter, R.D. & Cresswell, P. Impaired assembly and transport of HLA-A and -B antigens in a mutant TxB cell hybrid. EMBO J. 5, 943–949 (1986).

    CAS  Article  Google Scholar 

  7. 7

    Amit, S. et al. Axin-mediated CKI phosphorylation of β-catenin at Ser45: a molecular switch for the Wnt pathway. Genes Dev. 16, 1066–1076 (2002).

    CAS  Article  Google Scholar 

  8. 8

    Zhou, J., Dudley, M.E., Rosenberg, S.A. & Robbins, P.F. Persistence of multiple tumor-specific T-cell clones is associated with complete tumor regression in a melanoma patient receiving adoptive cell transfer therapy. J. Immunother. 28, 53–62 (2005).

    Article  Google Scholar 

  9. 9

    Goshima, G., Mayer, M., Zhang, N., Stuurman, N. & Vale, R.D. Augmin: a protein complex required for centrosome-independent microtubule generation within the spindle. J. Cell Biol. 181, 421–429 (2008).

    CAS  Article  Google Scholar 

  10. 10

    Rammensee, H., Bachmann, J., Emmerich, N.P., Bachor, O.A. & Stevanovic, S. SYFPEITHI: database for MHC ligands and peptide motifs. Immunogenetics 50, 213–219 (1999).

    CAS  Article  Google Scholar 

  11. 11

    Morel, S. et al. Processing of some antigens by the standard proteasome but not by the immunoproteasome results in poor presentation by dendritic cells. Immunity 12, 107–117 (2000).

    CAS  Article  Google Scholar 

  12. 12

    Chapiro, J. et al. Destructive cleavage of antigenic peptides either by the immunoproteasome or by the standard proteasome results in differential antigen presentation. J. Immunol. 176, 1053–1061 (2006).

    CAS  Article  Google Scholar 

  13. 13

    Guillaume, B. et al. Two abundant proteasome subtypes that uniquely process some antigens presented by HLA class I molecules. Proc. Natl. Acad. Sci. USA 107, 18599–18604 (2010).

    CAS  Article  Google Scholar 

  14. 14

    Kahle, J.J. et al. Comparison of an expanded ataxia interactome with patient medical records reveals a relationship between macular degeneration and ataxia. Hum. Mol. Genet. 20, 510–527 (2011).

    CAS  Article  Google Scholar 

  15. 15

    Blazek, D. et al. The cyclin K/Cdk12 complex maintains genomic stability via regulation of expression of DNA damage response genes. Genes Dev. 25, 2158–2172 (2011).

    CAS  Article  Google Scholar 

  16. 16

    Kawakami, Y. et al. Cloning of the gene coding for a shared human melanoma antigen recognized by autologous T cells infiltrating into tumor. Proc. Natl. Acad. Sci. USA 91, 3515–3519 (1994).

    CAS  Article  Google Scholar 

  17. 17

    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).

    CAS  Article  Google Scholar 

  18. 18

    Boël, P. et al. BAGE: a new gene encoding an antigen recognized on human melanomas by cytolytic T lymphocytes. Immunity 2, 167–175 (1995).

    Article  Google Scholar 

  19. 19

    Robbins, P.F. et al. A mutated β-catenin gene encodes a melanoma-specific antigen recognized by tumor infiltrating lymphocytes. J. Exp. Med. 183, 1185–1192 (1996).

    CAS  Article  Google Scholar 

  20. 20

    Cox, A.L. et al. Identification of a peptide recognized by five melanoma-specific human cytotoxic T cell lines. Science 264, 716–719 (1994).

    CAS  Article  Google Scholar 

  21. 21

    Pieper, R. et al. Biochemical identification of a mutated human melanoma antigen recognized by CD4+ T cells. J. Exp. Med. 189, 757–766 (1999).

    CAS  Article  Google Scholar 

  22. 22

    van der Bruggen, P., Stroobant, V., Vigneron, N. & Van den Eynde, B. Peptide database: T cell–defined tumor antigens. Cancer Immun.〉 (2013).

  23. 23

    Matsushita, H. et al. Cancer exome analysis reveals a T-cell–dependent mechanism of cancer immunoediting. Nature 482, 400–404 (2012).

    CAS  Article  Google Scholar 

  24. 24

    Castle, J.C. et al. Exploiting the mutanome for tumor vaccination. Cancer Res. 72, 1081–1091 (2012).

    CAS  Article  Google Scholar 

  25. 25

    Kvistborg, P. et al. TIL therapy broadens the tumor-reactive CD8+ T cell compartment in melanoma patients. OncoImmunology 1, 409–418 (2012).

    Article  Google Scholar 

  26. 26

    Jones, S. et al. Frequent mutations of chromatin remodeling gene ARID1A in ovarian clear cell carcinoma. Science 330, 228–231 (2010).

    CAS  Article  Google Scholar 

  27. 27

    Greenman, C. et al. Patterns of somatic mutation in human cancer genomes. Nature 446, 153–158 (2007).

    CAS  Article  Google Scholar 

  28. 28

    Berger, M.F. et al. Melanoma genome sequencing reveals frequent PREX2 mutations. Nature 485, 502–506 (2012).

    CAS  Article  Google Scholar 

  29. 29

    Topalian, S.L., Muul, L.M., Solomon, D. & Rosenberg, S.A. Expansion of human tumor infiltrating lymphocytes for use in immunotherapy trials. J. Immunol. Methods 102, 127–141 (1987).

    CAS  Article  Google Scholar 

  30. 30

    Dudley, M.E. et al. Adoptive cell transfer therapy following non-myeloablative but lymphodepleting chemotherapy for the treatment of patients with refractory metastatic melanoma. J. Clin. Oncol. 23, 2346–2357 (2005).

    CAS  Article  Google Scholar 

  31. 31

    Arnold, D. et al. Proteasome subunits encoded in the MHC are not generally required for the processing of peptides bound by MHC class I molecules. Nature 360, 171–174 (1992).

    CAS  Article  Google Scholar 

Download references


We thank B. Van den Eynde (Ludwig Institute) for providing HEK293 cells transfected with immunoproteasomal subunits and S. Schwarz and R. Fisch for assisting with experiments.

Author information




P.F.R. designed and developed the experimental screening system, analyzed data and drafted the manuscript. Y.-C.L. and M.E.-G. performed experiments evaluating TIL responses against candidate mutated peptides and analyzed results. Y.F.L. cloned and sequenced gene products encoding candidate epitopes identified by exomic sequencing and analyzed results. J.K.T., C.G., E.T., J.C.L. and P.C. carried out bioinformatic analyses. J.G. provided advice on exomic sequencing, prepared samples for sequencing and carried out validation studies using Sanger sequencing. Y.S. provided advice on sequencing of DNA isolated from tumor and normal cells and assisted with data analysis. S.A.R. helped design the studies and edited the manuscript.

Corresponding author

Correspondence to Paul F Robbins.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Tables 1–7 (PDF 213 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Robbins, P., Lu, YC., El-Gamil, M. et al. Mining exomic sequencing data to identify mutated antigens recognized by adoptively transferred tumor-reactive T cells. Nat Med 19, 747–752 (2013).

Download citation

Further reading


Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing