Inhibition of cellular NEDDylation partially alters the MHC class I immunopeptidome.

Abstract

Determining how chemotherapeutic drugs alter the MHC class I-associated immunopeptidome is essential for designing rational combinatorial therapies to enhance antitumor CD8+ T-cell activity, such as immune checkpoint inhibitor Abs. We investigated the impact of a first-in-class inhibitor of cellular NEDDylation, MLN4924, on self-peptide presentation using 2 human tumor cell lines: HeLa cells secreting the human MHC class I protein HLA-A*11:01 and the glioblastoma cell line U87-MG. MLN4924 indirectly inactivates the cullin-RING ligases (CRLs), a large family of E3 ubiquitin ligases responsible for targeting numerous substrates for degradation by the ubiquitin-proteasome system. MLN4924 treatment prevented cell division and altered the transcriptome of HeLa cells in a predictable manner. However, MLN4924 treatment did not alter the cell-surface levels of MHC class I proteins. An analysis of the immunopeptidome and the source proteins, which yielded peptides for presentation, revealed some unexpected findings. Greater than 50% of the peptides presented, and more than two-thirds of the source proteins from which peptides were derived, remained unaltered after MLN4924 treatment. Furthermore, alterations to the transcriptome could not predict the observed changes to the immunopeptidome. Of the approximately 100 CRL substrates that were presented by HLA-A*11:01, most were presented independent of MLN4924 treatment. Alternative degradation pathways and the presence of intron sequences in the specific mRNA transcripts may help explain how these proteins yield peptides for Ag presentation. In summary, MLN4924 treatment only partially affects the immunopeptidomic profile, suggesting it could be used in combinatorial cancer immunotherapies.