Elsevier

Vaccine

Volume 31, Issue 13, 25 March 2013, Pages 1725-1733
Vaccine

Immune enhancing properties of the novel Matrix-M™ adjuvant leads to potentiated immune responses to an influenza vaccine in mice

https://doi.org/10.1016/j.vaccine.2013.01.039Get rights and content

Abstract

The novel saponin based adjuvant Matrix-M™ was recently used in a Phase I study of seasonal influenza in elderly. The present study is a pre-clinical evaluation of the efficacy and mode-of-action of Matrix-M™ formulated influenza vaccine in mice. A manuscript on safety profile and immunogenicity in elderly humans is under preparation.

We have previously shown that subcutaneous injections of Matrix-M™, without coformulated antigen, results in a dose-dependent recruitment of leukocytes to draining lymph nodes (dLNs). Herein we compared the mode of action of Matrix-M™ with Alum, FCA and AS03 alone or formulated with influenza split virion antigen injected intramuscularly. The elicited responses in dLNs and spleen were investigated 48 h later. Matrix-M™ was particularly efficient in activation of central innate immune cells such as neutrophils, DCs and macrophages compared to the other adjuvants analyzed. Moreover, the adjuvant influence on the recall immune response to influenza antigen was studied by in vitro re-stimulation of splenocytes from mice immunized with influenza antigen adjuvanted with Matrix-M™, Alum or AS03. Splenocytes from mice immunized with influenza antigen and Matrix-M™ produced both Th1 and Th2 cytokines upon re-stimulation. This response was significantly stronger than that induced by the other adjuvants studied. Interestingly, increased levels of the neutrophil chemoattractant KC were produced by antigen stimulated splenocytes from mice immunized with Matrix-M™ adjuvanted vaccine, which is in agreement with the increase of neutrophils into dLNs and spleen after Matrix-M™ injection. Furthermore, influenza antigen adjuvanted with Matrix-M™ induced significantly higher antigen-specific IgG1 and IgG2a responses compared to antigen alone. In conclusion, adjuvant Matrix-M™ activates the innate immune system without antigen present. This activation may explain the enhanced immunity to influenza seen with Matrix-M™ adjuvant. Despite this potent immune activation mediated by Matrix-M™, GLP-toxicity studies and clinical data suggest that Matrix-M™ adjuvant has a mild to moderate safety profile.

Highlights

Mice received Matrix-M™, AS03, Alum or FCA, spleen, lymph nodes, and serum were analyzed. ► Immune response compared after vaccination with adjuvanted influenza antigen. ► Matrix-M™ activates innate immune cells, not seen by other investigated adjuvants. ► Splenocyte re-stimulation shows enhanced cytokine response in Matrix-M™ treated mice. ► High antibody levels in mice injected with Matrix-M™- or AS03-adjuvanted vaccine.

Introduction

Adjuvants that activate innate immune responses are thought to increase the likelihood of achieving efficient vaccine formulations. Many types of adjuvants potentiate immune responses by activating innate immune cells leading to cytokine secretion and cell recruitment, events important for antigen uptake and presentation. Saponins from the tree Quillaja saponaria Molina have for long been used as adjuvant in veterinary vaccines [1]. Matrix-M™ is a novel saponin-based adjuvant composed of purified saponin, cholesterol and phospholipids formulated into 40 nm cage-like structures. Particulate saponin adjuvants such as Matrix-M™ and immune stimulatory complexes (ISCOMs) are known to enhance cell trafficking, activate immune cells and have antigen dose-sparing potential [2], [3]. Furthermore they are known to induce Th1 and Th2 responses, i.e. to activate cytotoxic T lymphocytes (CTLs) alongside a strong humoral response [4], [5], [6]. Other adjuvants, such as Adjuvant System 03 (AS03), composed of squalene and α-tocopherol, also enhance Th1- and Th2-responses, demonstrated by increased IFN-γ production by in vitro re-stimulation alongside high hemagglutination inhibition titers after immunization with AS03-adjuvanted influenza [7]. In contrast, aluminum containing adjuvants (Alum) and MF59, based on squalene, are thought to mainly activate Th2 responses [8], [9]. Recently, it was shown that both Alum and MF59, in presence of antigen, recruit and activate neutrophils, monocytes and dendritic cells (DCs), cells important for antigen uptake and transportation to draining lymph nodes (dLNs) [10], [11], [12]. Thus antigen transportation may be a major function induced by Alum and MF59. Another widely used adjuvant in experimental research and commercial antibody manufacturing is Freund's Complete Adjuvant (FCA). FCA is composed of mineral oil mixed with killed mycobacteria, which creates a depot effect at the injection site. FCA enables efficient Th1 and Th2 responses [8]. Characterization of adjuvants’ mechanism of action will increase the knowledge on adjuvant design and efficient vaccine formulations for activation of both arms of the immune system. Hence, herein we have compared immune stimulatory properties of Matrix-M™, AS03, Alum and FCA without antigen present. In addition, Matrix-M formulated influenza antigen, identical to a formulation recently used in a Phase 1 study in elderly, was evaluated immunologically in mice. Humoral and cellular responses were compared to those induced by AS03 and Alum formulated with the same influenza antigen.

Section snippets

Adjuvants

Four adjuvants were studied: Matrix-M™ (AbISCO®-100, Isconova AB, Sweden) composed of two 40 nm large particles, i.e. Matrix-A™ and Matrix-C™, made from separate saponin fractions, Alhydrogel (Brenntag, Denmark) 2% Al(OH)3-suspension, FCA (Brenntag) formulated by mineral oil and heat-killed mycobacteria and AS03 (Pandemrix, GlaxoSmithKline) made of squalene and α-tocopherol.

Mice and injections

Female BALB/c mice (8–12 weeks) were purchased from, and kept at, the National Veterinary Institute (SVA, Sweden). All

Increased cellularity in dLNs and spleen after Matrix-M™ administration

To study the inherent potency of different adjuvants in recruitment and activation of immune cells during the early immune response, mice were injected s.c. with Matrix-M™, Alum, FCA or AS03 alone and the responses were compared to PBS-treated mice. After 48 h, dLNs from Matrix-M™-treated mice contained significantly higher cell numbers. This was not seen for Alum, FCA or AS03 (Fig. 1A). Also in spleen, Matrix-M™ induced a significant higher total cell count, though not as pronounced as for dLNs

Discussion

Adjuvant activities of Matrix-M™ were studied and compared to the following adjuvants: Alum, widely used and known to stimulate antibody responses in a Th2 biased manner [13], AS03, oil-based adjuvant with a balanced Th1/Th2 stimulatory effect with demonstrated efficacy in a human pandemic influenza vaccine [7], [14], FCA, inducing potent antibody and cellular responses [15]. Evaluation of inherent immune stimulatory properties by the adjuvants alone demonstrated significant differences between

References (23)

  • H. Takahashi et al.

    Induction of CD8+ cytotoxic T cells by immunization with purified HIV-1 envelope protein in ISCOMs

    Nature

    (1990)
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    Present address: Fiomi Diagnostics AB, Dag Hammarskjöldsväg 52A, SE-752 37 Uppsala, Sweden.

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