Heterologous Prime-Boost Combinations Highlight the Crucial Role of Adjuvant in Priming the Immune System

Abstract

The induction and modulation of the immune response to vaccination can be rationally designed by combining different vaccine formulations for priming and boosting. Here, we investigated the impact of heterologous prime-boost approaches on the vaccine-specific cellular and humoral responses specific for a mycobacterial vaccine antigen. C57BL/6 mice were primed with the chimeric vaccine antigen H56 administered alone or with the CAF01 adjuvant, and boosted with H56 alone, or combined with CAF01 or with the squalene-based oil-in-water emulsion adjuvant (o/w squalene). A strong secondary H56-specific CD4+ T cell response was recalled by all the booster vaccine formulations when mice had been primed with H56 and CAF01, but not with H56 alone. The polyfunctional nature of T helper cells was analyzed and visualized with the multidimensional flow cytometry FlowSOM software, implemented as a package of the R environment. A similar cytokine profile was detected in groups primed with H56 + CAF01 and boosted with or without adjuvant, except for some clusters of cells expressing high level of IL-17 together with TNF-α, IL-2, and IFN-γ, that were significantly upregulated only in groups boosted with the adjuvants. On the contrary, the comparison between groups primed with or without the adjuvant showed a completely different clusterization of cells, strengthening the impact of the formulation used for primary immunization on the profiling of responding cells. The presence of the CAF01 adjuvant in the priming formulation deeply affected also the secondary humoral response, especially in groups boosted with H56 alone or o/w squalene. In conclusion, the presence of CAF01 adjuvant in the primary immunization is crucial for promoting primary T and B cell responses that can be efficiently reactivated by booster immunization also performed with antigen alone.

Transcriptomics of the Vaccine Immune Response: Priming With Adjuvant Modulates Recall Innate Responses After Boosting

“…While we observed only a moderate differential expression of T cell activation modules 7 days after boost, the immunological analysis performed 10 days after boosting showed a higher frequency of H56-specific CD4+ T cells and germinal center B cells in dLN, a strong H56-specific humoral response and a higher frequency of ASC in spleen of mice primed with H56 + CAF01. We and others have previously shown that the presence of the CAF01 adjuvant combined with the antigen in the vaccine formulation used for priming is crucial for the induction of the germinal center reaction, assessed by the presence of the T follicular and germinal center B cells ( 4 , 46 ), as well as for a strong T cell mediated immune response with a Th1 and Th17 profile ( 4 , 6 , 47 ).…”

Computational Analysis of Multiparametric Flow Cytometric Data to Dissect B Cell Subsets in Vaccine Studies

“…The chimeric antigen H56, a promising vaccine candidate against Mycobacterium tuberculosis was used as model vaccine antigen, and administered by the parenteral route alone or combined with the CAF01 adjuvant, a liposome system capable to elicit both humoral and cellular responses . Boosting was performed with a lower dose of antigen alone, according to an immunization schedule already tested . The analysis was aimed to identify, in an automated and unbiased way, different B cell subtypes elicited by booster immunization and determine their frequency among mice primed with or without the adjuvant component.…”

“…FlowSOM is considered one of the best high‐performance algorithms in automated identification of cell subsets showing an extremely fast runtime . It has also been used for characterizing both the cell phenotype and the cellular functionality, such as the simultaneous production of intracellular cytokine and degranulation . The FlowSOM algorithm is based on a self‐organizing map (SOM), where similar cells are assigned to the same node.…”

“…Moreover, activation of B cells leads to the generation of subtypes of cells with different functions, such as plasmablasts, short‐or long‐lived plasma cells (PCs), memory cells . The presence of the adjuvant, especially in the primary immunization, allows to enhance and modulate the immune responses to vaccination and ensures the induction of long‐lived immunological memory necessary for protection .…”

From Bivariate to Multivariate Analysis of Cytometric Data: Overview of Computational Methods and Their Application in Vaccination Studies

“…Automated analysis can be particularly efficient also for identifying polyfunctional antigen-specific T cells elicited by vaccine administration or natural infection. Different computational tools, ranging from the Boolean combination gates, FlowSOM, or integrated approach combining targeted feature extraction (OpenCyto) with dimension reduction (t-SNE) have indeed been used to profile the polyfunctional activity of tuberculosis antigen-specific T cells and visualize treatment-specific differences between different vaccine formulations [106][107][108]. These studies demonstrate the importance of automated approaches to identify and visualize changes in very rare, multifunctional, antigen-specific T cells across different conditions, in flow cytometry datasets.…”