Bonanza of Spliced Epitopes to Enrich Immune Science

    Peptides displayed by the human leukocyte antigen (HLA) system have been subjected to unusually thorough prospecting. As a result, a mother lode of potential new targets has been uncovered for developers of immunotherapies and vaccines. These targets are spliced epitopes, merged peptides processed by antigen-presenting cells. Once thought rare, spliced epitopes now appear to account for a large proportion of all epitopes.

    In a new study, researchers based at Imperial College London and Charité–Universitätsmedizin Berlin have discovered that around one-third of all the epitopes displayed for scanning by the immune system are spliced epitopes. The scientists identified thousands of them by developing a new method that facilitates the mapping of cell surfaces.

    Details of the work appeared October 21 in the journal Science, in an article entitled, “A Large Fraction of HLA Class I Ligands Are Proteasome-Generated Spliced Peptides.” The article not only indicates that the proteasome-generated spliced peptide pool accounts for about one-third of the entire HLA class I immunopeptidome in terms of diversity, but about one-fourth of the immunopeptidome in terms of abundance as well.

    “This pool also represents a unique set of antigens, possessing particular and distinguishing features,” wrote the authors. “We validated this observation using a range of complementary experimental and bioinformatics approaches, as well as multiple cell types.”

    The article also describes an analytical strategy that the scientists used to account for recent discoveries underpinning the proteasome-catalyzed peptide splicing (PCPS) mechanism. With this strategy, which proved capable of handling the vast proteome-wide human spliced peptide database, the scientists undertook an initial analysis of the HLA-I–eluted immunopeptidome of the GR lymphoblastoid cell line. For a deeper coverage of the immunopeptidome, the scientists adopted a two-dimensional peptide prefractionation strategy followed by a hybrid peptide fragmentation method (electron-transfer higher-energy collision dissociation) for peptide identification, supplemented by an adapted target-decoy approach.

    “Our analysis led to the identification of 6,592 nonspliced and 3,417 spliced peptides 9 to 12 amino acid residues in length (9- to 12-mer peptides),” the scientists reported. “The latter number represents 34% of the total of identified antigenic peptides, thereby increasing the number of identified HLA-I ligands by some 50%.”

    These extra epitopes give the immune system more to scan and more possibilities of detecting disease. However, as the spliced epitopes are mixed sequences, they also have the potential to overlap with the sequences of healthy signalers and be misidentified as harmful.

    This could help scientists understand autoimmune diseases, where the immune system turns against normal body tissues, such as in type 1 diabetes and multiple sclerosis.

    The study’s lead author, Juliane Liepe from the Department of Life Sciences at Imperial, said: “The discovery of the importance of spliced peptides could present pros and cons when researching the immune system. For example, the discovery could influence new immunotherapies and vaccines by providing new target epitopes for boosting the immune system, but it also means we need to screen for many more epitopes when designing personalized medicine approaches.”

    The authors of the Science article concluded that the widespread appearance and abundance of proteasome-catalyzed peptide splicing events has implications for immunobiology and autoimmunity theories and may provide a previously untapped source of epitopes for use in vaccines and cancer immunotherapy.

    “It’s as if a geographer would tell you they had discovered a new continent, or an astronomer would say they had found a new planet in the solar system,” noted Michael Stumpf, a professor in the Department of Life Sciences at Imperial and a co-author of the study. “And just as with those discoveries, we have a lot of exploring to do. This could lead to not only a deeper understanding of how the immune system operates, but also suggests new avenues for therapies and drug and vaccine development.”