Powerful Complex Immunoadjuvant Based on Synergistic Effect of Combined TLR4 and NOD2 Activation Significantly Enhances Magnitude of Humoral and Cellular Adaptive Immune Responses

• Published in: PloS one Vol. 11; no. 5; p. e0155650
• Main Authors: Tukhvatulin, Amir I, Dzharullaeva, Alina S, Tukhvatulina, Natalia M, Shcheblyakov, Dmitry V, Shmarov, Maxim M, Dolzhikova, Inna V, Stanhope-Baker, Patricia, Naroditsky, Boris S, Gudkov, Andrei V, Logunov, Denis Y, Gintsburg, Alexander L
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 17.05.2016; Public Library of Science (PLoS)
• Subjects: Acetylmuramyl-Alanyl-Isoglutamine - immunology; Adaptive Immunity; Adjuvant chemotherapy; Adjuvants; Adjuvants, Immunologic; Alum; Aluminum sulfate; Analysis; Antibodies; Antigens; Antimicrobial peptides; Binding; Biology and Life Sciences; Bone marrow; CD4 antigen; CD8 antigen; CD80 antigen; CD86 antigen; Cell activation; Cell Line; Cell proliferation; Cell-mediated immunity; Cellular manufacture; Chemokines; Cholera; Cytokines; Dendritic cells; Drosophila; Epidemiology; Formulations; Gene expression; Humans; Immune response; Immune response (humoral); Immune system; Immunity; Immunity, Cellular; Immunity, Humoral; Immunization; Immunogenicity; Immunogenicity, Vaccine; Immunoglobulin G; Immunostimulation; Inflammation; Innate immunity; Insects; Lipid A - analogs & derivatives; Lipid A - immunology; Lymphocytes; Lymphocytes T; Medicine and Health Sciences; Nod2 Signaling Adaptor Protein - immunology; Oligomerization; Ovalbumin; Ovalbumin - immunology; Pattern recognition; Pattern recognition receptors; People and Places; Peptides; Proteins; Receptors; Receptors, Pattern Recognition - agonists; Receptors, Pattern Recognition - immunology; Spleen; Stimulation; Synergistic effect; T cell receptors; Th1 Cells - immunology; Toll-Like Receptor 4 - immunology; Toll-like receptors; Transcription; Vaccines; γ-Interferon; Cleveland Ohio; New York; United States > US; Russia
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0155650

Binding of pattern recognition receptors (PRRs) by pathogen-associated molecular patterns (PAMPs) activates innate immune responses and contributes to development of adaptive immunity. Simultaneous stimulation of different types of PRRs can have synergistic immunostimulatory effects resulting in enhanced production of molecules that mediate innate immunity such as inflammatory cytokines, antimicrobial peptides, etc. Here, we evaluated the impact of combined stimulation of PRRs from different families on adaptive immunity by generating alum-based vaccine formulations with ovalbumin as a model antigen and the Toll-like receptor 4 (TLR4) agonist MPLA and the Nucleotide-binding oligomerization domain-containing protein 2 (NOD2) agonist MDP adsorbed individually or together on the alum-ovalbumin particles. Multiple in vitro and in vivo readouts of immune system activation all showed that while individual PRR agonists increased the immunogenicity of vaccines compared to alum alone, the combination of both PRR agonists was significantly more effective. Combined stimulation of TLR4 and NOD2 results in a stronger and broader transcriptional response in THP-1 cells compared to individual PRR stimulation. Immunostimulatory composition containing both PRR agonists (MPLA and MDP) in the context of the alum-based ovalbumin vaccine also enhanced uptake of vaccine particles by bone marrow derived dendritic cells (BMDCs) and promoted maturation (up-regulation of expression of CD80, CD86, MHCII) and activation (production of cytokines) of BMDCs. Finally, immunization of mice with vaccine particles containing both PRR agonists resulted in enhanced cellular immunity as indicated by increased proliferation and activation (IFN-γ production) of splenic CD4+ and CD8+ T cells following in vitro restimulation with ovalbumin and enhanced humoral immunity as indicated by higher titers of ovalbumin-specific IgG antibodies. These results indicate that combined stimulation of TLR4 and NOD2 receptors dramatically enhances activation of both the humoral and cellular branches of adaptive immunity and suggests that inclusion of agonists of these receptors in standard alum-based adjuvants could be used to improve the effectiveness of vaccination.