Molecular and Cellular Response Profiles Induced by the TLR4 Agonist-Based Adjuvant Glucopyranosyl Lipid A

Abstract

BackgroundToll-like receptor (TLR)4 agonists are known potent immunostimulatory compounds. These compounds can be formulated as part of novel adjuvants to enhance vaccine medicated immune responses. However, the contribution of the formulation to the innate in vivo activity of TLR4 agonist compounds is not well understood.Methodology and Principal FindingsWe evaluated synthetic TLR4 agonist Glucopyranosyl Lipid A (GLA) for its effects on molecular and cellular innate immune responses in the murine model. Microarray techniques were used to compare the responses to GLA in an aqueous formulation or in an oil-in-water Stable Emulsion formulation (GLA-SE) versus either SE alone or the mineral salt aluminum hydroxide (alum) at the muscle injection site over multiple timepoints. In contrast to the minimal gene upregulation induced by SE and alum, both GLA and GLA-SE triggered MyD88- and TRIF-dependent gene expression. Genes for chemokines, cytokine receptors, signaling molecules, complement, and antigen presentation were also strongly upregulated by GLA and GLA-SE. These included chemokines for TH1-type T cells (CXCL9 and CXCL10) and mononuclear leukocytes (CCL2, CCL3) among others. GLA-SE induced stronger and more sustained gene upregulation than GLA in the muscle; GLA-SE induced genes were also detected in local draining lymph nodes and at lower levels in peripheral blood. Both GLA and GLA-SE resulted in increased cellular trafficking to the draining lymph nodes and upregulated MHC molecules and ICAM1 on local dendritic cells. GLA and GLA-SE transiently upregulated circulating MCP-1, TNFα, IFNγ and IP-10 in blood.Conclusions/SignificanceWhile GLA and GLA-SE activate a large number of shared innate genes and proteins, GLA-SE induces a quantitatively and qualitatively stronger response than GLA, SE or alum. The genes and proteins upregulated could be used to facilitate selection of appropriate adjuvant doses in vaccine formulations.

Molecular signatures of vaccine adjuvants

“…Transcriptional profiles induced by aqueous GLA and GLA-SE, including Cxcl9, Cxcl10, Cxcl11, Ifng, Stat1 and Ccr5 as well as Th1-associated serum cytokines CCL2, CXCL1, TNF␣, IL-6, IFN-␥ and CSF3 corroborated the documented Th1-baising property of these adjuvants [30]. Genes involved in MyD88 and TRIF dependent pathways were also up regulated by both GLA-SE and GLA [29]. This is in line with a recent finding in which the Th1 polarizing effect of GLA-SE was shown to be dependent on both MyD88 and TRIF signaling pathways [28].…”

“…GLA-SE has been tested in clinical trials as part of exploratory vaccines against tuberculosis, malaria, HIV and other diseases [28]. Transcriptome profiling of the site of injection, draining lymph nodes and blood at several early time points provided important information on the kinetics and magnitude of gene expression induced by GLA-SE [29]. The overall duration and magnitude of gene expression at the injection site, cytokine responses in serum as well as leucocyte recruitment to the muscle and the draining lymph nodes were more profound after GLA-SE administration (peaking at 48 h) compared to those of the aqueous GLA formulation (peaking at 6 h) whereas SE alone (peaking at 96 h) induced only limited gene expression.…”

Current advancements and potential strategies in the development of MERS-CoV vaccines

“…Normally, vaccines with limited immunogenicity require the addition of specific adjuvants to stimulate a protective and long-lasting immune response [96–98]. Although a variety of novel adjuvants are now emerging and being investigated, only a few have been approved for clinical use.…”

Impact of formulation and particle size on stability and immunogenicity of oil-in-water emulsion adjuvants

“…25 To this end, the concentration of adjuvant-induced protein cytokines IL-6, MCP-1, KC, and IP-10 were evaluated in mouse sera 6 h post immunization. In the absence of CpG 2395, the expression level of the cytokines appeared to be inversely correlated to the particle size (see Table 2 for correlation coefficients), with the highest induction observed in the animals immunized with the 80 nm particles (Fig.…”

“…These emulsions were then placed in stability chambers at 5 C, 25 C, 40 C, and 60 C and monitored over the course of a month. In addition, samples were also placed on an orbital shaker at 120 rpm (at 25 C) to assess the impact on shear stress on the emulsion stability.…”