PXD070374

PXD070374 is an original dataset announced via ProteomeXchange.

Title	Raising the Bar for Isobaric Amino Acid Variants in Immunopeptidome of Pancreatic Cancer: Distinguishing Ile-Leu variant Ligands by ExCID
Description	Immunopeptidomics aims to facilitate T-cell target discovery by analyzing human leukocyte antigen (HLA)-bound peptides using tandem mass spectrometry (MS/MS). MS/MS analysis of peptides conventionally relies on collision-induced dissociation (CID) of selected precursor ions. However, CID analysis of immunopeptides, owing to their diverse termini (non-tryptic) and relatively short lengths (8- to 12-mer for class I HLA), can be challenging. It is impossible to differentiate isobaric amino acids, such as isoleucine and leucine sequence variants, which are frequent amino acids in the immunopeptidomes of many HLA allotypes. To address this, we leveraged the alternative fragmentation method, Electron-Activated Dissociation (EAD), and its CID-supplemented mode (EAD+CID, or ExCID) on the SCIEX ZenoTOF 7600+ system to characterize Ile/Leu sequence variants in the immunopeptidome. We first used a panel of seven synthetic Ile/Leu-substituted peptides and demonstrated that both EAD and ExCID efficiently generate variant-distinguishing w- and d-type satellite ions. We then determined optimal kinetic energy (KE) and EAD reaction time for utilizing hybrid ExCID ion activation in a discovery immunopeptidomics setting. We demonstrated that Ile/Leu sequence variants present in the immunopeptidome can be distinguished using ExCID but not CID alone. Using this optimized ExCID methodology, we observed improvements in fragmentation and peptide identification for both HLA class I and II studies, allowing us to benefit from EAD while minimizing the compromise to cycle time and identification depth. We further applied the optimal ExCID method to complex HLA peptide samples isolated from two model cell lines, namely a “monoallelic” HLA-A2 expressing cell line (C1R-A*02:01) and the pancreatic cancer cell line Panc1. The versatile and tuneable hybrid ExCID mode improves MS/MS fragment coverage and enriches the product spectra with diagnostic ions, enabling us to differentiate and characterize Ile/Leu sequence variants in immunopeptidomics studies.
HostingRepository	PRIDE
AnnounceDate	2026-06-16
AnnouncementXML	Submission_2026-06-16_00:16:06.013.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Mohammadreza Dorvash
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: NEWT:9606;
ModificationList	phosphorylated residue; acetylated residue; monohydroxylated residue; deamidated residue
Instrument	ZenoTOF 7600+

Revision	Datetime	Status	ChangeLog Entry
0	2025-11-05 12:06:52	ID requested
⏵ 1	2026-06-16 00:16:06	announced

10.1021/ACS.ANALCHEM.5C08123;

10.1021/acs.analchem.5c08123;

submitter keyword: None

Professor Anthony
contact affiliation	Department of Biochemistry and Molecular Biology and the Biomedicine Discovery Institute; Monash University; Clayton 3800; Victoria; Australia
contact email	anthony.purcell@monash.edu
lab head
Mohammadreza Dorvash
contact affiliation	Department of Biochemistry and Molecular Biology and the Biomedicine Discovery Institute; Monash University; Clayton 3800; Victoria; Australia
contact email	mohammadreza.dorvash@monash.edu
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD070374

PRIDE project URI

• PRIDE
  1. PXD070374
     1. Label: PRIDE project
     2. Name: Raising the Bar for Isobaric Amino Acid Variants in Immunopeptidome of Pancreatic Cancer: Distinguishing Ile-Leu variant Ligands by ExCID