Pharmaceutical Research

, Volume 34, Issue 8, pp 1716–1727 | Cite as

Systematic Investigation of the Role of Surfactant Composition and Choice of oil: Design of a Nanoemulsion-Based Adjuvant Inducing Concomitant Humoral and CD4+ T-Cell Responses

  • Signe Tandrup Schmidt
  • Malene Aaby Neustrup
  • Stine Harloff-Helleberg
  • Karen Smith Korsholm
  • Thomas Rades
  • Peter Andersen
  • Dennis Christensen
  • Camilla Foged
Research Paper
  • 215 Downloads

Abstract

Purpose

Induction of cell-mediated immune (CMI) responses is crucial for vaccine-mediated protection against difficult vaccine targets, e.g., Chlamydia trachomatis (Ct). Adjuvants are included in subunit vaccines to potentiate immune responses, but many marketed adjuvants stimulate predominantly humoral immune responses. Therefore, there is an unmet medical need for new adjuvants, which potentiate humoral and CMI responses. The purpose was to design an oil-in-water nanoemulsion adjuvant containing a synthetic CMI-inducing mycobacterial monomycoloyl glycerol (MMG) analogue to concomitantly induce humoral and CMI responses.

Methods

The influence of emulsion composition was analyzed using a systematic approach. Three factors were varied: i) saturation of the oil phase, ii) type and saturation of the applied surfactant mixture, and iii) surfactant mixture net charge.

Results

The emulsions were colloidally stable with a droplet diameter of 150–250 nm, and the zeta-potential correlated closely with the net charge of the surfactant mixture. Only cationic emulsions containing the unsaturated surfactant mixture induced concomitant humoral and CMI responses upon immunization of mice with a Ct antigen, and the responses were enhanced when squalene was applied as the oil phase. In contrast, emulsions with neutral and net negative zeta-potentials did not induce CMI responses. The saturation degree of the oil phase did not influence the adjuvanticity.

Conclusion

Cationic, MMG analogue-containing nanoemulsions are potential adjuvants for vaccines against pathogens for which both humoral and CMI responses are needed.

Key Words

adjuvant drug delivery emulsion immune response vaccine 

Abbreviations

CAF

Cationic adjuvant formulation

CMI

Cell-mediated immunity

Cryo-TEM

Cryo-transmission electron microscopy

Ct

Chlamydia trachomatis

DDA

Dimethyldioctadecylammonium bromide

DODAC

Dioleoyldimethylammonium chloride

DOPE

Dioleoylphosphoethanolamine

DSPE

Distearoylphosphoethanolamine

GLA

Glucopyranosyl lipid A

GRAS

Generally regarded as safe

HA

Hemagglutinin

HEPES

4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid

HLB

Hydrophile-lipophile balance

HRP

Horseradish peroxidase

HSM

High shear mixing

IFN

Interferon

IL

Interleukin

LN

Lymph node

MHC

Major histocompatibility complex

MMG-1

Monomycoloyl glycerol

MOMP

Major outer membrane protein

Mtb

Mycobacterium tuberculosis

o/w

Oil-in-water

PDI

Polydispersity index

PMA

Phorbol-12-myristate-13-acetate

Pos:neu

Positive-to-neutral

RSV

Respiratory syncytial virus

rt.

Room temperature

S.c.

Subcutaneous

TDB

Trehalose-6,6′-dibehenate

TIV

Trivalent influenza vaccine

TMB

3,3′,5,5′-tetramethylbenzidine

Tris

Tris(hydroxymethyl)aminomethane

w/o

Water-in-oil

z-average

Intensity-weighted average hydrodynamic diameter

Supplementary material

11095_2017_2180_MOESM1_ESM.pptx (104 kb)
Supplementary data, Figure 1Gating strategy for the assessment of cytokine producing T cells by using flow cytometry. Single cells were further gated into lymphocytes, from which CD4+ T-cell were identified. The expression of IFN-γ, IL-17a, IL-2 and TNF-α was subsequently determined. The sample show splenocytes from a mouse immunized with CAF04-adjuvanted OVA, which were restimulated with CTH522. (PPTX 104 kb)
11095_2017_2180_MOESM2_ESM.pptx (87 kb)
Supplementary data, Figure 2Hydrodynamic diameter of emulsion droplets measured during optimization of process parameters for high pressure homogenization, performed on emulsions composed of squalene, Span 60 and Tween 60. The z-average of the pre-emulsion was approx. 530 nm (data not shown). Data points represent mean values ± SD (n = 1, three technical replicates). (PPTX 86 kb)

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Copyright information

© Springer Science+Business Media New York 2017

Authors and Affiliations

  • Signe Tandrup Schmidt
    • 1
    • 2
  • Malene Aaby Neustrup
    • 1
  • Stine Harloff-Helleberg
    • 1
  • Karen Smith Korsholm
    • 2
  • Thomas Rades
    • 1
  • Peter Andersen
    • 2
  • Dennis Christensen
    • 2
  • Camilla Foged
    • 1
  1. 1.Department of Pharmacy, Faculty of Health and Medical SciencesUniversity of CopenhagenCopenhagen ØDenmark
  2. 2.Statens Serum Institut, Department of Infectious Disease ImmunologyCopenhagen SDenmark

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