Preprint Club
A cross-institutional Journal Club Initiative

Keywords

• Mycobacterium tuberculosis

• HLA-II peptides

• Mass spectrometry

• Proteome-wide discovery

• Reverse vaccinology

Main Findings

Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB), leads to millions of deaths every year, thereby surpassing other human infectious diseases. Upon infection, individuals develop a cell-mediated immune response driven by CD4+ T cells. Yet, this response does not always clear the microbial agent, which may eventually progress to active TB disease. To protect infants and children from life-threatening TB, the bacillus Calmette-Guérin (BCG) vaccine obtained from Mycobacterium bovis has been used worldwide. By contrast, the vaccination outcome in adults is rather low. Thus, to control the global TB epidemic, newer vaccines with substantial power to fuel endogenous T-cell responses are urgently needed. Aiming to fill this knowledge/clinical gap, Leddy et al. show in this preprint (not peer-reviewed) a rapid and optimised state-of-the-art protocol based on mass spectrometry to discover immunogenic human leukocyte antigen class II (HLA-II)-binding peptides to design TB vaccines.

Immunopeptidomics robustly identifies immunogenic HLA-II peptides from elemental Mtb proteins.

Using Mtb-infected human monocyte-derived dendritic cells (hMDCs), the authors identified 27 peptides from 13 Mtb proteins, including EsxG, TatA, and EsxB. The latter, interestingly, has been already used as a main target in tested vaccine candidates. Further, membrane-associated Mtb antigens from the type VII secretion systems (e.g., PE/PPE proteins) commonly popped up in the HLA-II repertoire, suggesting that these antigens could be more accessible to the host HLA-II processing machinery. Of note, most of the HLA-II peptides identified by mass spectrometry triggered the production of IFNγ and IL-17 in peripheral blood mononuclear cells from individuals with prior Mtb exposure. Thus, this proteome-wide discovery approach offers new additional immunogenic targets that could be prioritised during the development of TB vaccines.

HLA-II peptides were highly conserved across the Mtb genomic diversity.

Notably, substitutions in the identified HLA-II binding peptides were positioned in the final branches of the phylogeny in all major Mtb lineages, affecting only < 0.15% of 51,229 Mtb isolates. This suggests that the Mtb diversity is unlikely to hinder vaccines targeting those HLA-II peptides. Moreover, when assessing the allele frequency spectrum of non-synonymous mutations in HLA-II peptides, the authors observed that Mtb strains harbouring nucleotide mutations in HLA-II peptides have less offspring, indicating that the identified immunogens are under purifying selection and, therefore, are not naturally removed as occurs in deleterious alleles.

A novel method to rapidly assess whether vaccine candidates may end up in HLA-II presentation.

Based on a successful method (SureQuant) previously employed to develop COVID-19 vaccines, the authors also set out a protocol to examine the ability of vaccines to result in HLA-II presentation of target antigens without using HLA-transgenic mice or thousands of volunteers—a relevant improvement that could simplify and shorten expensive resources before the onset of early clinical trials. This procedure allowed them to assess whether the BCG vaccine could lead to HLA-II presentation of the same peptides displayed by human Mtb-infected cells. Importantly, the data show for the first time that both BCG- and Mtb-infected cells present conserved HLA-II peptides from EsxG, PE19, and PE5/PE29 antigens.

Quantitative targeted mass spectrometry is suitable to optimise mRNA vaccines against microbial agents.

So far, the powerful mRNA vaccine technology has essentially targeted viral infections, whereas the development of mRNA vaccines to fight intracellular microbial infections are yet to be optimised. Of note, Leddy et al. present a major refinement in countermeasure against Mtb by designing and directing mRNA-encoded immunogens to a variety of subcellular compartments (mitochondria, endoplasmic reticulum, endosomes, and lysosomes) to enhance HLA-II presentation. This revealed that the HLA-II presentation of mRNA-encoded immunogens was more efficient when directing them to lysosomes.

Hence, this research represents a pioneer dataset exploring the nature of HLA-II peptides from Mtb in infected human cells, thus offering potent immunogens to accelerate the development of a new generation of TB vaccines.

Limitations

• As required during the development of any drug or vaccine candidate, a thorough investigation of the toxicity and allergenicity of the identified HLA-II peptides as well as the assessment of potential adjuvants would provide more fundamental insights to translate these findings into clinical applications.

• The in silico modelling of the three-dimensional structure of microbial peptides followed by their docking with the most common HLA-II alleles in the world's population would aid in understanding: 1) remarkable molecular patterns during the interaction between Mtb epitopes and HLA-II molecules, 2) suitability of other vaccine platforms such as peptide vaccines conjugated and formulated as nanoparticles, and 3) whether the vaccine construct(s) could achieve a successful global immunisation coverage.

Significance/Novelty

TB is a serious infectious disease that affects a quarter of the global population and threatens to spread to new regions as climate change spikes. Hence, the development and optimisation of TB vaccines is a fundamental goal in the field. Leddy et al. provide an innovative, robust, and rapid perspective to harness reverse vaccinology by cutting-edge mass spectrometry and thus to discover conserved and highly immunogenic Mtb epitopes to develop mRNA TB vaccines. For instance, instead of using antigens that harbour fewer immunogenic peptides, a more potent vaccine can be designed by identifying and selecting the most immunogenic targets to boost the CD4+ T cell response. Remarkably, the implications of translating these findings to fight other severe diseases, including cancer, are quite promising.

Credit

Reviewed by Jose G. Marchan-Álvarez as part of a cross-institutional journal club between the Icahn School of Medicine at Mount Sinai, the University of Oxford, the Karolinska Institute, the University of Toronto and the University of Texas MD Aderson Cancer Center.

The author declares no conflict of interests in relation to their involvement in the review.