Biological responses induced aluminum-based vaccine adjuvants

Abstract

Vaccination is, together with clean drinking water, one of the most important means to prevent the spread and to control infectious diseases. Since the introduction of vaccines in the 18th century, the mortality rate caused by infectious diseases has drastically decreased. In 1926, Alexander Thomas Glenny discovered that a vaccine containing aluminum salts provided better protection towards the disease than a vaccine with only the pathogen. Since that time aluminum salts are often used in vaccines as adjuvants. A comprehensive understanding of the working mechanism of adjuvants, however, is still missing. This knowledge is necessary to modify and improve vaccine-induced immune responses. The research described in this thesis reveals the effect of defined adjuvants on cells involved in the initiation phase of the immune response, e.g. monocytes both at a molecular level (transcriptome and proteome) as well as the biological process in and between cells. By combining various techniques, including flow cytometry, ELISA, qPCR and mass spectrometry, a comprehensive overview of the processes activated by these adjuvants was generated. It was observed that Al(OH)3 (one of the aluminum-based adjuvants) prepares monocytes for the induction of an immune response even in the absence of an antigen. Additionally, it was observed that Al(OH)3 itself is capable of inducing cytokine secretion related to an inflammatory response and Th2 polarization. The response towards this adjuvant is significantly altered by antigens in a vaccine: for example, the antigens in DTaP (Diphtheria, Tetanus and acellular Pertussis) impacted the vaccine response both qualitatively and quantitatively, since a Th1 inhibiting cytokine was observed upon DTaP stimulation but not upon Al(OH)3 stimulation. Moreover, the inflammatory cytokine IL-b was induced much stronger by DTaP. The immune responses towards different aluminum-based adjuvants are not the same: Al(OH)3 stimulation of mice resulted in a neutrophil influx at the site of injection, while this was not the case for AlPO4. Size as well as shape of the aluminum salt particles also influence the immune response. The immune response towards the nanoparticle-sized Al(OH)3 adjuvant was substantially weaker compared to the response towards the microparticle of the Al(OH)3 adjuvant. In addition, needle-shaped adjuvants induced a much stronger response than the hexagon-shaped product. An improved mass spectrometric-based assay was also developed to profile the protein secretion of de novo-synthesized proteins after a stimulation (e.g. an adjuvant). This method is based on cyclooctyne-functionalized agarose beads which allows for a rapid and catalyst-free capture of proteins containing azido-moieties incorporated in the proteins as methionine analogues. The advantage of cyclooctyne-functionalized beads is that the click chemistry does not require any catalyst and that this assay does not require a preselection of the targeted cytokines/chemokines as is the case in Multiplex Immunoassays (e.g. Luminex). The research described in this thesis shows that the biological responses towards aluminum-based adjuvants are very complex. Hence, it is essential to test different adjuvant-antigen combinations already in an early phase of the vaccine development and optimization.