IMBAS-MS Discovers Organ-Specific HLA Peptide Patterns in Plasma

摘要

•Automated one-pot enrichment of soluble HLA peptides using magnetic beads (IMBAS).•Personalized computational library generation for DIA analysis using AlphaPeptDeep.•IMBAS-MS identifies more than 5000 HLA peptides form 200 μl of plasma.•Soluble HLA peptides represent a variety of tissues including the brain. Distinction of non-self from self is the major task of the immune system. Immunopeptidomics studies the peptide repertoire presented by the human leukocyte antigen (HLA) protein, usually on tissues. However, HLA peptides are also bound to plasma soluble HLA (sHLA), but little is known about their origin and potential for biomarker discovery in this readily available biofluid. Currently, immunopeptidomics is hampered by complex workflows and limited sensitivity, typically requiring several mL of plasma. Here, we take advantage of recent improvements in the throughput and sensitivity of mass spectrometry (MS)-based proteomics to develop a highly sensitive, automated, and economical workflow for HLA peptide analysis, termed Immunopeptidomics by Biotinylated Antibodies and Streptavidin (IMBAS). IMBAS-MS quantifies more than 5000 HLA class I peptides from only 200 μl of plasma, in just 30 min. Our technology revealed that the plasma immunopeptidome of healthy donors is remarkably stable throughout the year and strongly correlated between individuals with overlapping HLA types. Immunopeptides originating from diverse tissues, including the brain, are proportionately represented. We conclude that sHLAs are a promising avenue for immunology and potentially for precision oncology. Distinction of non-self from self is the major task of the immune system. Immunopeptidomics studies the peptide repertoire presented by the human leukocyte antigen (HLA) protein, usually on tissues. However, HLA peptides are also bound to plasma soluble HLA (sHLA), but little is known about their origin and potential for biomarker discovery in this readily available biofluid. Currently, immunopeptidomics is hampered by complex workflows and limited sensitivity, typically requiring several mL of plasma. Here, we take advantage of recent improvements in the throughput and sensitivity of mass spectrometry (MS)-based proteomics to develop a highly sensitive, automated, and economical workflow for HLA peptide analysis, termed Immunopeptidomics by Biotinylated Antibodies and Streptavidin (IMBAS). IMBAS-MS quantifies more than 5000 HLA class I peptides from only 200 μl of plasma, in just 30 min. Our technology revealed that the plasma immunopeptidome of healthy donors is remarkably stable throughout the year and strongly correlated between individuals with overlapping HLA types. Immunopeptides originating from diverse tissues, including the brain, are proportionately represented. We conclude that sHLAs are a promising avenue for immunology and potentially for precision oncology. The immune system relies on the human leukocyte antigen (HLA) peptide-protein complex to present immunopeptides derived from both endogenous and exogenous sources to circulating T-cells. These immunopeptides play a crucial role in immune surveillance, as they enable the elimination of abnormal or infected cells. Upon recognition of a peptide–HLA protein complex by cytotoxic T lymphocytes (CTLs), downstream cascades are activated, causing the presenting cell to undergo apoptosis. This biological principle is exploited in immunotherapeutic strategies such as CAR T-cell treatment or mRNA peptide vaccination (1Nelde A. Maringer Y. Bilich T. Salih H.R. Roerden M. Heitmann J.S. et al.Immunopeptidomics-guided warehouse design for peptide-based immunotherapy in chronic lymphocytic leukemia.Front. Immunol. 2021; 12: 705974Crossref PubMed Scopus (17) Google Scholar). A crucial but challenging step is the identification of peptides specifically presented by tumor cells. Most efforts have focused on the enrichment of membrane-bound human leukocyte antigen (mHLA) receptors with their bound immunope