Clin Exp Vaccine Res.  2015 Jan;4(1):88-98. 10.7774/cevr.2015.4.1.88.

Systemic antibody response to nano-size calcium phospate biocompatible adjuvant adsorbed HEV-71 killed vaccine

• Saeed MI
• Omar AR
• Hussein MZ
• Elkhidir I
• Sekawi Z

• KMID: 2049112
• DOI: http://doi.org/10.7774/cevr.2015.4.1.88

Abstract

PURPOSE
Since 1980s, human enterovirus-71 virus (HEV-71) is one of the common infectious disease in Asian Pacific region since late 1970s without effective commercial antiviral or protective vaccine is unavailable yet. The work examines the role of vaccine adjuvant particle size and the route of administration on postvaccination antibody response towards HEV-71 vaccine adsorbed to calcium phosphate (CaP) adjuvant.
MATERIALS AND METHODS
First, CaP nano-particles were compared to a commercial micro-size and vaccine alone. Secondly, intradermal reduced dosage was compared to the conventional intramuscular immunization. Killed HEV-71 vaccines adsorbed to CaP nano-size (73 nm) and commercial one of micro-size (1.7 microm) were administered through intradermal, intramuscular, rabbits received vaccine alone and unvaccinated animals.
RESULTS
CaP nano-particles adsorbed HEV-71 vaccine displayed higher antibody than the micro-size or unadsorbed vaccine alone, through both parenteral immunization routes. Moreover, the intradermal route (0.5 microg/mL) of 0.1-mL volume per vaccine dose induced equal IgG antibody level to 1.0-mL intramuscular route (0.5 microg/mL).
CONCLUSION
The intradermal vaccine adsorbed CaP nano-adjuvant showed safer and significant antibody response after one-tenth reduced dose quantity (0.5 microg/mL) of only 0.1-mL volume as the most suitable protective, cost effective and affordable formulation not only for HEV-71; but also for developing further effective vaccines toward other human pathogens.

Keyword

Human enterovirus 71; Calcium phosphate; Adjuvanted vaccine; Nanoparticles; Intradermal injections

MeSH Terms

Animals
Antibody Formation*
Asian Continental Ancestry Group
Calcium*
Communicable Diseases
Enterovirus A, Human
Humans
Immunization
Immunoglobulin G
Injections, Intradermal
Nanoparticles
Particle Size
Rabbits
Vaccines
Calcium
Immunoglobulin G
Vaccines

Figure

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  Fig. 1 The mechanism of calcium phosphate biocompatible adjuvant of a non-immunogenic advantage over other adjuvants.

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  Fig. 2 Inactivated virus showed normal monolayer of Vero cell (A) and human enterovirus-71 virus-positive control showed viral cytopathic effect at 36-hour postinfection (B).

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  Fig. 3 Scanning electron microscope. (A) Calcium phosphate (CaP) nano-particles (×100,000). (B) Commercial CaP adjuvant micro-particle, 1.7 µm (×25,000). (C) Transmission electron microscope micrograph. Human enterovirus-71 virus of 31 nm (×100,000).

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  Fig. 4 (A) The nano-prepared calcim phosphate (CaP nano-particles) adjuvant of mean particles size, 30 nm. (B) Commercial adjuvant (CaP micro-praticles) of mean particle size, 905 nm.

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  Fig. 5 Adjuvants cytotoxicity; both did not have adverse effect on human liver cell proliferation, due the biocompatibility of the calcium phosphate adjuvant.

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  Fig. 6 IgM antibodies were higher in rabbits immunized with vaccine adsorbed nano-calcium phosphate adjuvant through intramuscular (IM) (A), while initially micro-adjuvant (red) showed increased level which dropped faster than the nano-adjuvant after the third dose of vaccine (B). HEV-71, human enterovirus-71 virus; ID, intradermal.

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  Fig. 7 IgG level was elevated in rabbits immunized intramuscular (IM) with nano-adjuvant adsorbed vaccine compared to vaccine alone (A). While IgG level was higher in rabbits immunized intradermal (ID) with nano-adjuvant adsorbed vaccine compared less in micro-adjuvant adsorbed and it was the least in animal received vaccine alone (B). HEV-71, human enterovirus-71 virus.

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  Fig. 8 IgG levels were higher vaccine adsorbed nano-adjuvant administered through intramuscular (IM) 1.0-mL volume (0.5 µg/mL), then nano-adjuvant administered intradermal (ID) only 0.1-mL volume (0.5 µg/mL) nano-adsorbed vaccine and less with ID 0.1-mL volume (0.5 µg/mL) micro-size adsorbed vaccine throughout the last 6 weeks (3-9 weeks).

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  Fig. 9 IgG levels were high in intramuscular (IM) administered unadsorbed vaccine (0.5 µg/mL) of 1.0-mL dose volume, compared to the reduced (0.5 µg/mL) intradermal (ID) immunization of only 0.1-mL volume, which indicated the role of using both adjuvant adsorbed vaccine and in a nano-size particles.

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  Fig. 10 Human enterovirus-71 virus (HEV-71) postvaccination in vitro neutralizing mean antibody titer in rabbits serum: 1, Vaccine in nano-calcium phosphate (CaP) adjuvant (intradermal [ID], 1:1,600); 2, Vaccine micro-CaP adjuvant (ID, 1:800); 3, Unadsorbed vaccine (ID, 1:200); 4, Vaccine in nano-CaP adjuvant (intramuscular [IM], 1:1,600); 5, Unadsorbed vaccine (IM, 1:400).

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