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Mass spectrometric analysis of the HLA class I peptidome of melanoma cell lines as a promising tool for the identification of putative tumor-associated HLA epitopes


Melanoma is one of the most immunogenic tumors, and extensive lists of potential tumor rejection antigens have been collected during the last decades. By isolating human leukocyte antigen (HLA) class I complexes from five melanoma cell lines (FM-82, FM-93/2, Mel-624, MeWo and SK-Mel-5) and sequencing HLA-eluted peptides by mass spectrometry, we identified over 10,000 unique peptides with high confidence. The majority of the peptides were 8–11 amino acids in length and were predicted to bind to the respective HLA alleles. Over 250 epitopes, corresponding to previously described tumor-associated antigens, were identified, suggesting that HLA peptidome analysis may facilitate the characterization of putative tumor rejection antigens. MeWo and SK-Mel-5 cell lines were further interrogated for neo-epitopes, revealing one peptide from MeWo cells carrying an amino acid mutation. We also observed a remarkable overlap between A*03:01 peptides eluted from Mel-624 cells and A*03:01 peptides recovered from soluble HLA complexes purified from two melanoma patients, shedding light on the similarity of the HLA peptidome in cell lines and in patient-derived material. The reliable characterization of the HLA class I peptidome in melanoma promises to facilitate the identification of tumor rejection antigens and the development of immunotherapeutic strategies.

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American Type Culture Collection


Catalogue of Somatic Mutations in Cancer


European Searchable Tumour Line Database

ETH Zurich:

Eidgenössische Technische Hochschule Zürich


False discovery rate


Fast protein liquid chromatography


Higher-energy collisional dissociation


Human leukocyte antigen

IC50 :

Half maximal inhibitory concentration


Liquid chromatography–mass spectrometry


Mass spectrometry


Fragment mass spectra


Phenylmethylsulfonyl fluoride


Soluble human leukocyte antigen


Ultra high performance liquid chromatography


Wild type


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We thank Camilla Bacci (Philogen SpA) for providing W6/32 antibody.


This work was supported financially by ETH Zürich, the Swiss National Science Foundation, the European Union’s Seventh Framework Programme (FP7/2007-2013) under Grant Agreement Nos. 305309 (PRIAT) and 305608 (EURenOmics), the Bonveda Foundation and the European Research Council (ERC advanced Grant “ZAUBERKUGEL”).

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Correspondence to Tim Fugmann or Dario Neri.

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Dario Neri is co-founder of Philogen, shareholder and member of the board. Tim Fugmann and Danilo Ritz are employees of Philochem AG. The authors declare no additional conflict of interest.

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Gloger, A., Ritz, D., Fugmann, T. et al. Mass spectrometric analysis of the HLA class I peptidome of melanoma cell lines as a promising tool for the identification of putative tumor-associated HLA epitopes. Cancer Immunol Immunother 65, 1377–1393 (2016).

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