title: MHC-I Ligand Discovery Using Targeted Database Searches of Mass Spectrometry Data: Implications for T-Cell Immunotherapies.
abstract: Class I major histocompatibility complex (MHC-I)-bound peptide ligands dictate the activation and specificity of CD8+T cells and thus are important for devising T-cell immunotherapies. In recent times, advances in mass spectrometry (MS) have enabled the precise identification of these MHC-I peptides, wherein MS spectra are compared against a reference proteome. Unfortunately, matching these spectra to reference proteome databases is hindered by inflated search spaces attributed to a lack of enzyme restriction in the searches, limiting the efficiency with which MHC ligands are discovered. Here we offer a solution to this problem whereby we developed a targeted database search approach and accompanying tool SpectMHC, that is based on a priori-predicted MHC-I peptides. We first validated the approach using MS data from two different allotype-specific immunoprecipitates for the C57BL/6 mouse background. We then developed allotype-specific HLA databases to search previously published MS data sets of human peripheral blood mononuclear cells (PBMCs). This targeted search strategy improved peptide identifications for both mouse and human ligandomes by greater than 2-fold and is superior to traditional "no enzyme" searches of reference proteomes. Our targeted database search promises to uncover otherwise missed novel T-cell epitopes of therapeutic potential.
authors.affiliation: Department of Microbiology and Immunology, Dalhousie University , Halifax, Nova Scotia B3H 4R2, Canada.
authors.forename: J Patrick
authors.initials: JP
authors.lastname: Murphy
authors.affiliation: Department of Microbiology and Immunology, Dalhousie University , Halifax, Nova Scotia B3H 4R2, Canada.
authors.forename: Prathyusha
authors.initials: P
authors.lastname: Konda
authors.affiliation: Department of Immunology, Interfaculty Institute for Cell Biology, University of Tübingen , 72076 Tübingen, Germany.|Immatics Biotechnologies GmbH , 72076 Tübingen, Germany.
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Kowalewski
authors.affiliation: Department of Immunology, Interfaculty Institute for Cell Biology, University of Tübingen , 72076 Tübingen, Germany.|Immatics Biotechnologies GmbH , 72076 Tübingen, Germany.
authors.forename: Heiko
authors.initials: H
authors.lastname: Schuster
authors.affiliation: Department of Pathology, Dalhousie University , Halifax, Nova Scotia B3H 4R2, Canada.
authors.forename: Derek
authors.initials: D
authors.lastname: Clements
authors.affiliation: Department of Pathology, Dalhousie University , Halifax, Nova Scotia B3H 4R2, Canada.
authors.forename: Youra
authors.initials: Y
authors.lastname: Kim
authors.affiliation: Proteomics Core Facility, Dalhousie University , Halifax, Nova Scotia B3H 4R2, Canada.
authors.forename: Alejandro M
authors.initials: AM
authors.lastname: Cohen
authors.affiliation: Department of Microbiology and Immunology, Dalhousie University , Halifax, Nova Scotia B3H 4R2, Canada.
authors.forename: Tanveer
authors.initials: T
authors.lastname: Sharif
authors.affiliation: Department of Bio and Health Informatics, Technical University of Denmark , 2800 Kongens Lyngby, Denmark.
authors.forename: Morten
authors.initials: M
authors.lastname: Nielsen
authors.affiliation: Department of Immunology, Interfaculty Institute for Cell Biology, University of Tübingen , 72076 Tübingen, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Stevanovic
authors.affiliation: Department of Microbiology and Immunology, Dalhousie University , Halifax, Nova Scotia B3H 4R2, Canada.|Department of Pathology, Dalhousie University , Halifax, Nova Scotia B3H 4R2, Canada.
authors.forename: Patrick W
authors.initials: PW
authors.lastname: Lee
authors.affiliation: Department of Microbiology and Immunology, Dalhousie University , Halifax, Nova Scotia B3H 4R2, Canada.|Department of Pathology, Dalhousie University , Halifax, Nova Scotia B3H 4R2, Canada.|Centre for Innovative and Collaborative Health Services Research, IWK Health Centre , Halifax, Nova Scotia B3K 6R8, Canada.
authors.forename: Shashi
authors.identifier: 0000-0002-5427-0829
authors.initials: S
authors.lastname: Gujar
mesh_terms: D000818:Animals; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D018984:Epitopes, T-Lymphocyte / Q000235:genetics / Q000276:immunology*; D015395:Histocompatibility Antigens Class I / Q000235:genetics / Q000276:immunology; D006801:Humans; D007167:Immunotherapy; D008024:Ligands; D013058:Mass Spectrometry / Q000379:methods*; D051379:Mice; D010455:Peptides / Q000235:genetics / Q000276:immunology*
chemical_list: D018984:Epitopes, T-Lymphocyte; D015395:Histocompatibility Antigens Class I; D008024:Ligands; D010455:Peptides
keywords: CD8 T-cell epitopes; MHC ligandome; database searching; immunotherapy; mass spectrometry