Systems vaccinology and big data in the vaccine development chain

Raeven, René H. M.; van Riet, Elly; Meiring, Hugo D.; Metz, Bernard; Kersten, Gideon F. A.

Secreted proteins play an essential role in communication between immune cells. The immune secretome consists of a wide range of cytokines such as tumor necrosis factors, interferons and interleukins. Multiplex immunoassays and ELISA methods target predefined proteins from the secretome, based on cytokine-specific antibodies. In contrast, newly developed MS-based proteomics methods like BioOrthogonal Non-Canonical Amino acid Tagging (BONCAT) can target the full repertoire of secreted proteins, although limited to in vitro experiments only. In a BONCAT assay, an amino acid is replaced by an azido-containing analogue (e.g. methionine replacement by azidohomoalanine). The azido moiety allows for purification of excreted, low abundance, newly synthesized proteins from cell culture medium by click- chemistry using alkyne-functionalized beads. 38, 39 The amino acid modification does not affect the structure and function of the protein, nor does it alter the protein synthesis rate or cell viability. 40, 41 In combination with multiplexing technologies based on Tandem Mass Tagging (TMT), N,N-dimethyl leucine (DiLeu) tagging or neutron-encoded reagents, up to 25 samples can be multiplexed in a single run, for unbiased identification and relative quantification of de novo-synthesized proteins in the secretome. 42, 43 Limitations of MS-based proteomics lay in the fact that, like transcriptomics, it is dependent on the availability and quality of annotated protein databases, e.g. number of annotated genes or proteins. The sensitivity of the current generation of mass spectrometers is high, but could improve further to reach the levels achieved with, for example, quantitative polymerase chain reaction. However, as described here and by others, 44 MS is currently very useful for the characterization of a wide range of immunological mechanisms in the context of infection and vaccination.