Immunopeptidomics-based design of mRNA vaccine formulations against Listeria monocytogenes

Listeria monocytogenes (further referred to as Listeria) is a foodborne pathogen that can cause listeriosis, which can be dangerous specially among vulnerable individuals including immunocompromised individuals, elderly people, and pregnant women. Infection typically occurs by consumption of contaminated food such as contaminated dairy. The bacterium’s ability to grow even at refrigerator temperatures renders it a considerable risk factor in food industry. After ingestion, it can cross the intestinal barrier, enter blood stream, spread to liver and spleen, and possibly cross the blood–brain barrier or the fetoplacental barrier to inflict various complications.

Listeria is capable of hiding from the immune system by invading host cells. Once inside the endocytic vesicle, Listeria secretes phospholipase as well as the pore-forming toxin listeriolysin O to access the cytosol. Then it replicates and spreads to neighbouring cells. While human listeriosis is not a very common type of infection, it has a high fatality rate of up to 30% and unfortunately number of cases are increasing. For example, in 2017, 2502 cases of listeriosis were confirmed in the EU/EEA countries, while in 2007 only 1635 cases had been reported. Although cases of listeriosis are rising, no vaccines for Listeria are currently. So far mostly live attenuated, inactivated Listeria or bacterial ghost vaccines have been explored as preclinical vaccine candidates.

To extend the antigen knowledge on Listeria, this study applied an immunopeptidomics study on two infected human epithelial cell lines. Among more than 15,000 (15,767 to be exact) human self-peptides, the researchers could identify 68 Listeria immunopeptides from 42 different bacterial antigens. Along with several known antigens, many novel ones were detected, often derived from the bacterial periphery.

How was PEAKS used?

Mass spectrometry raw data were searched against database using PEAKS Studio X + (version 10.5). Database was of the human sequences in UniProt SwissProt (January 2019, with 20,413 entries) merged with Listeria monocytogenes EGD sequences from TrEMBL (April 2019, with 2847 entries).

The peptide length range was 8–30 amino acids, using unspecific digestion as digest mode. Methionine oxidation and N-terminal acetylation were set as variable modifications. Mass error tolerances were set to 10 ppm and 0.02 Da for parent and fragment ions, respectively. To check for potential contaminant peptides, the MaxQuant contaminant database (MQ version 1.6.3.4) was enabled. Identified peptide sequences were filtered at the PSM level for an FDR of 1% and higher prior to label-free or TMT-10plex quantification. Quantification results were not filtered, and TMT-labelled peptide quantifications were not normalized before exporting.

Mayer, Rupert L., et al. “Immunopeptidomics-based design of mRNA vaccine formulations against Listeria monocytogenes.” Nature Communications 13.1 (2022): 1-17. https://doi.org/10.1038/s41467-022-33721-y

Abstract

Listeria monocytogenes is a foodborne intracellular bacterial pathogen leading to human listeriosis. Despite a high mortality rate and increasing antibiotic resistance no clinically approved vaccine against Listeria is available. Attenuated Listeria strains offer protection and are tested as antitumor vaccine vectors, but would benefit from a better knowledge on immunodominant vector antigens. To identify novel antigens, we screen for Listeria peptides presented on the surface of infected human cell lines by mass spectrometry-based immunopeptidomics. In between more than 15,000 human self-peptides, we detect 68 Listeria immunopeptides from 42 different bacterial proteins, including several known antigens. Peptides presented on different cell lines are often derived from the same bacterial surface proteins, classifying these antigens as potential vaccine candidates. Encoding these highly presented antigens in lipid nanoparticle mRNA vaccine formulations results in specific CD8+ T-cell responses and induces protection in vaccination challenge experiments in mice. Our results can serve as a starting point for the development of a clinical mRNA vaccine against Listeria and aid to improve attenuated Listeria vaccines and vectors, demonstrating the power of immunopeptidomics for next-generation bacterial vaccine development.

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