SILAC-based quantification reveals modulation of the immunopeptidome in BRAF and MEK inhibitor sensitive and resistant melanoma cells
peer-reviewed · Frontiers in Immunology · 2025
| Date | 2025-01-06 |
| Type | peer-reviewed |
| Venue | Frontiers in Immunology |
| Publisher | Frontiers Media SA |
| Contribution | downstream-application |
| DOI | 10.3389/fimmu.2024.1490821 |
| Citations (OpenAlex) | 2 |
Abstract
Background The immunopeptidome is constantly monitored by T cells to detect foreign or aberrant HLA peptides. It is highly dynamic and reflects the current cellular state, enabling the immune system to recognize abnormal cellular conditions, such as those present in cancer cells. To precisely determine how changes in cellular processes, such as those induced by drug treatment, affect the immunopeptidome, quantitative immunopeptidomics approaches are essential. Methods To meet this need, we developed a pulsed SILAC-based method for quantitative immunopeptidomics. Metabolic labeling with lysine, arginine, and leucine enabled isotopic labeling of nearly all HLA peptides across all allotypes (> 90% on average). We established a data analysis workflow that integrates the de novo sequencing-based tool Peptide-PRISM for comprehensive HLA peptide identification with MaxQuant for accurate quantification. Results We employed this strategy to explore the modulation of the immunopeptidome upon MAPK pathway inhibition (MAPKi) and to investigate alterations associated with early cellular responses to inhibitor treatment and acquired resistance to MAPKi. Our analyses demonstrated significant changes in the immunopeptidome early during MAPKi treatment and in the resistant state. Moreover, we identified putative tumor-specific cryptic HLA peptides linked to these processes that might represent exploitable targets for cancer immunotherapy. Conclusions We have developed a new mass spectrometric approach that allowed us to investigate the effects of common MAPK inhibitors on the immunopeptidome of melanoma cells. This finally led to the discovery of new potential targets for cancer immunotherapy.
Methods and tools
- pSILAC quantitative immunopeptidomics with Peptide-PRISM: Pulsed SILAC labelling combined with Peptide-PRISM de novo-based identification and MaxQuant quantification tracks how MAPK inhibition reshapes the melanoma immunopeptidome, revealing cryptic HLA peptides.
Methods it uses
- PEAKS: Commercial DP-based de novo
- Peptide-PRISM: Identifies cryptic HLA-I peptides by matching the top de novo sequencing candidates for each spectrum against a six-frame translation of the genome and transcriptome, then scoring them in a stratified FDR; showed cryptic peptides are a substantial part of the HLA-I immunopeptidome.
Data deposited
- SILAC-based quantification reveals modulation of the immunopeptidome in BRAF and MEK inhibitor-treated and resistant tumor cells — as deposited · PXD054097