Identification of the Cryptic HLA-I Immunopeptidome.

Florian Erhard,Lars Dölken,Bastian Schilling,Andreas Schlosser

doi:10.1158/2326-6066.cir-19-0886

Florian Erhard, Lars Dölken + Show 2 more

Open Access

https://doi.org/10.1158/2326-6066.cir-19-0886

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Abstract

The success of cancer immunotherapy relies on the ability of cytotoxic T cells to specifically recognize and eliminate tumor cells based on peptides presented by HLA-I. Although the peptide epitopes that elicit the corresponding immune response often remain unidentified, it is generally assumed that neoantigens, due to tumor-specific mutations, are the most common targets. Here, we used a mass spectrometric approach to show an underappreciated class of epitopes that accounts for up to 15% of HLA-I peptides for certain HLA alleles in various tumors and patients. These peptides are translated from cryptic open reading frames in supposedly noncoding regions in the genome and are mostly unidentifiable with conventional computational analyses of mass spectrometry (MS) data. Our approach, Peptide-PRISM, identified thousands of such cryptic peptides in tumor immunopeptidomes. About 20% of these HLA-I peptides represented the C-terminus of the corresponding translation product, suggesting frequent proteasome-independent processing. Our data also revealed HLA-I allele-dependent presentation of cryptic peptides, with HLA-A*03 and HLA-A*11 presenting the highest percentage of cryptic peptides. Our analyses refute the reported frequent presentation of HLA peptides generated by proteasome-catalyzed peptide splicing. Thus, Peptide-PRISM represents an important step toward comprehensive identification of HLA-I immunopeptidomes and reveals cryptic peptides as an abundant class of epitopes with potential relevance for novel immunotherapeutic approaches.

Highlights

Immunotherapeutic approaches, such as adoptive T-cell therapy or therapeutic peptide vaccination, are among the most promising approaches to treat cancer [1]
By analyzing HLA-I immunopeptidome data, we previously found that at least 2% of the peptides bound to HLA-I in human fibroblasts originate from cryptic open reading frames (ORF) identified by Ribo-seq [13]
The tumor-specific antigens that are recognized by the corresponding cytotoxic T cells mediating the antitumor effect are mostly unknown

Summary

Introduction

Immunotherapeutic approaches, such as adoptive T-cell therapy or therapeutic peptide vaccination, are among the most promising approaches to treat cancer [1]. In spite of overwhelming evidence for their translation, only a limited number of their translation products have been detected by MS so far, even when adopted isolation methods and database search approaches have been applied [5, 6] This indicates that cryptic translation products are short-lived, but are exploited as a source for HLA-I peptides, as has been described for defective ribosomal products [7,8,9,10]. We present a fundamentally different strategy, termed Peptide-PRISM, to identify cryptic peptides based on mass spectrometric data alone This enabled us to analyze a large collection of HLA immunopeptidomes without using additional sequencing data, and to identify 6,636 cryptic peptides from various types of tumors from several patients with high confidence. This reveals cryptic HLA peptides to be a substantial part of tumor immunopeptidomes

Results

Discussion

Disclosure of Potential Conflicts of Interest

Erhard