No abstract

TLR ligands are used in modern vaccine adjuvants, TLR4 ligand-based adjuvants are the most advanced in commercial vaccines. Increased understanding of TLR4 receptor-ligand interactions enables chemical synthesis and modification of new leads and our understanding of the biological/immunological mechanisms of combination adjuvants enables formulation of potent and safe vaccine compositions. Characterization of non-glycolipid TLR4 ligands provided new mechanistic information that could lead to new formulations. This review discusses advances in TLR4 agonist design-both glycolipid and non-glycolipid based TLR4 ligands-as well as CD14 activation as options to activate or synergize with TLR4 signaling. Finally, we review the molecular and cellular mechanisms that are elicited by formulated TLR4 targeted combination adjuvants during the initiation of innate immune responses leading to quality adaptive responses.

Elderly people are at high risk for influenza-related morbidity and mortality due to progressive immunosenescence. While toll-like receptor (TLR) agonist containing adjuvants, and other adjuvants, have been shown to enhance influenza vaccine-induced protective responses, the mechanisms underlying how these adjuvanted vaccines could benefit the elderly remain elusive. Here, we show that a split H1N1 influenza vaccine (sH1N1) combined with a TLR4 agonist, glucopyranosyl lipid adjuvant formulated in a stable oil-in-water emulsion (GLA-SE), boosts IgG2c:IgG1 ratios, enhances hemagglutination inhibition (HAI) titers, and increases protection in aged mice. We find that all adjuvanted sH1N1 vaccines tested were able to protect both young and aged mice from lethal A/H1N1/California/4/2009 virus challenge after two immunizations compared to vaccine alone. We show that GLA-SE combined with sH1N1, however, also provides enhanced protection from morbidity in aged mice given one immunization (based on change in weight percentage). While the GLA-SE-adjuvanted sH1N1 vaccine promotes the generation of cytokine-producing T helper 1 cells, germinal center B cells, and long-lived bone marrow plasma cells in young mice, these responses were muted in aged mice. Differential in vitro responses, dependent on age, were also observed from mouse-derived bone marrow-derived dendritic cells and lung homogenates following stimulation with adjuvants, including GLA-SE. Besides enhanced HAI titers, additional protective factors elicited with sH1N1 + GLA-SE in young mice were observed, including (a) rapid reduction of viral titers in the lung, (b) prevention of excessive lung inflammation, and (c) homeostatic maintenance of alveolar macrophages (AMs) following H1N1 infection. Collectively, our results provide insight into mechanisms of adjuvant-mediated immune protection in the young and elderly.

Mycobacterium tuberculosis (M.tb), a bacterial pathogen that causes tuberculosis disease (TB), exerts an extensive burden on global health. The complex nature of M.tb, coupled with different TB disease stages, has made identifying immune correlates of protection challenging and subsequently slowing vaccine candidate progress. In this work, we leveraged two delivery platforms as prophylactic vaccines to assess immunity and subsequent efficacy against low-dose and ultra-low-dose aerosol challenges with M.tb H37Rv in C57BL/6 mice. Our second-generation TB vaccine candidate ID91 was produced as a fusion protein formulated with a synthetic TLR4 agonist (glucopyranosyl lipid adjuvant in a stable emulsion) or as a novel replicating-RNA (repRNA) formulated in a nanostructured lipid carrier. Protein subunit- and RNA-based vaccines preferentially elicit cellular immune responses to different ID91 epitopes. In a single prophylactic immunization screen, both platforms reduced pulmonary bacterial burden compared to the controls. Excitingly, in prime-boost strategies, the groups that received heterologous RNA-prime, protein-boost or combination immunizations demonstrated the greatest reduction in bacterial burden and a unique humoral and cellular immune response profile. These data are the first to report that repRNA platforms are a viable system for TB vaccines and should be pursued with high-priority M.tb antigens containing CD4+ and CD8+ T-cell epitopes.

Newly generated T cells must undergo phenotypic and functional maturation. Previously, we showed that the absence of NKAP severely curtailed T cell maturation, and NKAP-deficient recent thymic emigrants (RTEs) fail to enter the long-lived naïve peripheral T cell pool. The paucity of peripheral T cells in CD4-cre NKAP conditional knockout (cKO) mouse is not due to apoptosis as neither loss of Bax nor Bcl-2 or Bcl-xL transgenes rescued the maturation defect. Here, we show that CD55, a complement regulatory protein functioning to disassemble C3 convertases, plays a role in T cell maturation. The expression of CD55, but not Crry or CD59, increases concurrently with T cell maturation by two orders of magnitude. NKAP-deficient peripheral T cells display a diminished expression of CD55 and an increase in complement C3 deposition, indicating CD55 is required for peripheral T cell fitness of RTEs. Reconstitution of CD4-cre NKAP cKO bone marrow into C3-deficient host partially rescues the maturation defect as measured by upregulation of Qa2 and CD55, and an increased proportion of mature naïve T cells. Thus, successful T cell maturation results in increased expression of CD55 that protects RTEs from complement-mediated killing.