Background: Antigen-presenting cells play a crucial role in directing T-cells to identify and eliminate cancer cells by presenting peptides on the cell surface in complex with Human Leukocyte Antigens (HLA). The peptides presented by HLA class I molecules originate from proteins that are degraded by the proteasome into linear and spliced (non-contiguous sequences derived from the same or different proteins) peptide antigens. Mapping the peptide ligands presented to the immune system – not only on cancerous tissue but also on cells within the tumour microenvironment – will greatly enhance the range and diversity of targets available for immuno-therapy and enable personalised cancer treatment.
Methods: This study combines peptide sequencing performed by mass spectrometry with PEAKS Studio software and novel bioinformatics algorithms to identify linear and spliced peptides presented by six patient-derived melanoma cells under normal and interferon-treated conditions.
Results: We created a database consisting of more than 30,000 unique peptides per cell line, including linear, cis- and trans-spliced peptides. Spliced peptides made up around 15-20% of each peptide dataset. Interferon treatment of cells dramatically remodelled the repertoire of peptides presented by all cell lines, with only ~40% overlap between peptides presented under constitutive and stimulated conditions. 3265 (~8% of the total dataset) presented by the HLA-A*02:01 allele were common to all cell lines. This database also includes nearly 1300 peptides derived from 79 known melanoma-associated antigens (MAA) and across all cell lines, about 50% of peptides derived from MAA were spliced in nature.
Conclusion: This study created the largest known database of melanoma patient-derived immunopeptidomes. We observed that interferon treatment leads to a change in the peptide repertoire presented by HLA-I and that peptide splicing further diversifies the immunopeptidome. A subset of HLA-A*02:01 binders were common to all cell lines, offering potential as viable vaccine targets across populations.