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.
Highlights
We employed proteomics to characterize the adjuvant-induced changes of secretomes from human newborn and adult monocytes in response to aluminum phosphate (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
The number of proteins identified in supernatants from differentially treated cells ranged from 829 to 1184 for the neonatal, and from 832 to 1406 for the adult samples, respectively. supplemental Table S1 gives an overview of these proteins, which were identified in seven biological replicates combined, with a raw spectral count Ն 1 and a protein false discovery rate (FDR) Ͻ 1%
Age alone appeared to exert an effect on TLR agonist (TLRA)-induced secretomes, resulting in distinct newborn and adult clusters. Both the TLRAs MPLA (TLR4) and R848 (TLR8), induced similar secretome clusters by age that were highly distinct from those induced by Alum (Fig. 2C)
Summary
When compared with their adult counterparts in vitro, human newborn and infant leukocytes demonstrate reduced Th1-polarizing cytokine induction in response to most TLRAs, including the TLR4As LPS and MPLA [2] For this reason, we choose to study human monocytes, leukocytes with antigen-presenting properties that are key players in the vaccine-induced immune response [18, 19]. We found that [1] adjuvant-induced monocyte secretomes varied by adjuvant and age, [2] that adjuvants induce distinct innate immune pathways, [3] that protein biomarkers defined by this approach were confirmed in adjuvant and licensed adjuvanted vaccine-stimulated whole blood assays in vitro as well as in [4] publicly available data sets of individuals receiving a novel adjuvanted vaccine in vivo. Our observations provide fresh insight into adjuvant action and suggest a novel paradigm for characterizing age-specific adjuvant effects in vitro in relation to their effects in vivo
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