Adjuvant-induced Human Monocyte Secretome Profiles Reveal Adjuvant- and Age-specific Protein Signatures

Abstract

Adjuvants boost vaccine responses, enhancing protective immunity against infections that are most common among the very young. Many adjuvants activate innate immunity, some via Toll-Like Receptors (TLRs), whose activities varies with age. Accordingly, characterization of age-specific adjuvant-induced immune responses may inform rational adjuvant design targeting vulnerable populations. In this study, we employed proteomics to characterize the adjuvant-induced changes of secretomes from human newborn and adult monocytes in response to Alum, the most commonly used adjuvant in licensed vaccines; Monophosphoryl Lipid A (MPLA), a TLR4-activating adjuvant component of a licensed Human Papilloma Virus vaccine; and R848 an imidazoquinoline TLR7/8 agonist that is a candidate adjuvant for early life vaccines. Monocytes were incubated in vitro for 24 h with vehicle, Alum, MPLA, or R848 and supernatants collected for proteomic analysis employing liquid chromatography-mass spectrometry (LC-MS) (data available via ProteomeXchange, ID PXD003534). 1894 non-redundant proteins were identified, of which ∼30 - 40% were common to all treatment conditions and ∼5% were treatment-specific. Adjuvant-stimulated secretome profiles, as identified by cluster analyses of over-represented proteins, varied with age and adjuvant type. Adjuvants, especially Alum, activated multiple innate immune pathways as assessed by functional enrichment analyses. Release of lactoferrin, pentraxin 3, and matrix metalloproteinase-9 was confirmed in newborn and adult whole blood and blood monocytes stimulated with adjuvants alone or adjuvanted licensed vaccines with distinct clinical reactogenicity profiles. MPLA-induced adult monocyte secretome profiles correlated in silico with transcriptome profiles induced in adults immunized with the MPLA-adjuvanted RTS,S malaria vaccine (Mosquirix™). Overall, adjuvants such as Alum, MPLA and R848 give rise to distinct and age-specific monocyte secretome profiles, paralleling responses to adjuvant-containing vaccines in vivo. Age-specific in vitro modeling coupled with proteomics may provide fresh insight into the ontogeny of adjuvant action thereby informing targeted adjuvanted vaccine development for distinct age groups.

Harnessing the beneficial heterologous effects of vaccination

“…Human newborn and adult monocytes exhibit distinct responses to adjuvants and licensed adjuvanted vaccines in vitro that demonstrate age- and adjuvant-specific correlations to adjuvanted vaccine-induced responses in vivo 99 . Moreover, TLR agonist administration to newborn mice enhances their subsequent cytokine and phagocytic responses to polymicrobial sepsis 100 , and early inflammatory/innate immune-activating events in humans are associated with reduced risk of late-onset sepsis 82, 83 .…”

A Meningococcal Outer Membrane Vesicle Vaccine Incorporating Genetically Attenuated Endotoxin Dissociates Inflammation from Immunogenicity

“…We have previously demonstrated that several of these licensed vaccines, such as the EasyFive (DTwP-HepB-Hib), induce distinct reactogenicity biomarker profiles in vitro (31, 32). When tested at equivalent v/v treatment concentrations, Δ lpxLI nOMV conversely induced a lower cytokine response for most innate cytokines (Figures 4A,B) and chemokines (Figure S7 in Supplementary Material) tested, grouping closer to pediatric vaccines such as PCV13 and HBV, than the more inflammatory (Alum + TLR agonist)-containing pediatric vaccines (i.e., PedvaxHIB and EasyFive).…”

“…Next, immature newborn (Figure 5A) and adult DCs (Figure 5B) were assessed for vaccine-induced production of PGE 2 , a molecule whose in vitro production has been correlated with reactogenicity in vivo (31–33). Of note, newborn DCs demonstrated significantly reduced Δ lpxLI nOMV-mediated PGE 2 responses as compared to both Bexsero ( p  < 0.01) and WT dOMV (with Alum) ( p  < 0.05) (Figure 5A).…”

“…Therefore, further study of the IL-1β/PGE 2 axis as a predictive marker of vaccine reactogenicity may need to take ontogeny into account. Recent use of proteomics (secretomics) has profiled hundreds of proteins released by human monocytes when stimulated in vitro with Alum or TLR agonists, several of which have been validated upon study of licensed adjuvanted vaccines, providing opportunities for further refinement of future adjuvanticity and reactogenicity biomarkers (31). Finally, the evaluation of different pentaacyl lipid A mutants (46) and comparison of their modified agonist properties using age-specific human WB and DC assays should also considered.…”

Adjuvant pretreatment with alum protects neonatal mice in sepsis through myeloid cell activation

“…However, it altered the phenotype of the recruited cells, including increased activation status, phagocytosis and NET formation, with a significant reduction in the bacterial colonization. Recently, the global proteomic response of human neonatal monocytes to alum demonstrated the activation of phagocytosis-related signalling (caveolarmediated endocytosis, MSP-RON) when monocytes were stimulated with alum in vitro [34]. Professional phagocytic cells such as macrophages are able to sense alum crystals in vitro via the activation of the NLRP3 inflammasome [35].…”