Calcium Phosphate Nanoparticles Induce Mucosal Immunity and Protection against Herpes Simplex Virus Type 2

Abstract

Previously we reported that calcium phosphate nanoparticles (CAP) represented a superior alternative to alum adjuvants in mice immunized with viral protein. Additionally, we showed that CAP was safe and elicited no detectable immunoglobulin E (IgE) response. In this study, we demonstrated that following mucosal delivery of herpes simplex virus type 2 (HSV-2) antigen with CAP, CAP adjuvant enhanced protective systemic and mucosal immunity versus live virus. Mice were immunized intravaginally and intranasally with HSV-2 protein plus CAP adjuvant (HSV-2؉CAP), CAP alone, phosphate-buffered saline, or HSV-2 alone. HSV-2؉CAP induced HSV-specific mucosal IgA and IgG and concurrently enhanced systemic IgG responses. Our results demonstrate the potency of CAP as a mucosal adjuvant. Furthermore, we show that systemic immunity could be induced via the mucosal route following inoculation with CAP-based vaccine. Moreover, neutralizing antibodies were found in the sera of mice immunized intranasally or intravaginally with HSV-2؉CAP. Also, the results of our in vivo experiments indicated that mice vaccinated with HSV-2؉CAP were protected against live HSV-2 infection. In conclusion, these preclinical data support the hypothesis that CAP may be an effective mucosal adjuvant that protects against viral infection.Since mucosal surfaces act as the primary point of entry for most pathogens and the first line of defense against them, vaccines inducing effective mucosal immunity may reduce rates of infection and decrease the morbidity and mortality of infectious diseases. Currently, no safe and effective mucosal vaccine adjuvants are approved for human use.Mucosal vaccine delivery is a promising strategy. Mucosal vaccines administered in one part of the body can elicit an antibody response in mucosal tissues remote from the site of initial antigen exposure. This effect occurs because of the common mucosal immune system (13). A major obstacle to developing a mucosal vaccine in humans is finding a safe and effective adjuvant. Experimental mucosal adjuvants include cholera toxin, heat-labile enterotoxin, mutant toxins (LTK63 and LTR72), CpG oligodeoxynucleotide, polymerized liposomes, microparticles, and interleukins or immune modulators. None of these adjuvants is approved for use in humans (3,12,18).Biodegradable calcium phosphate particles have been investigated as an alternative to aluminum adjuvants for parenteral vaccines. Clinical studies conducted in France described the use of a calcium phosphate adjuvant for secondary or booster immunizations against diphtheria and tetanus (8). Calcium phosphate has also been used for allergen desensitization (6, 16). Early studies indicated that calcium phosphate particles produce strong adjuvant effects, induced less immunoglobulin E (IgE) than aluminum adjuvants, and elicited only minimal local irritation in animal experiments and human clinical trials (6,8,11,17).Here, we describe a unique formulation of calcium phosphate nanoparticles (CAP) which is distinct from the formulations of calciu...

Development of a nasal vaccine for chronic hepatitis B infection that uses the ability of hepatitis B core antigen to stimulate a strong Th1 response against hepatitis B surface antigen

“…The ELISPOT assay using the P815 mastocytome cell line pulsed with the CD8 + -restricted HBs (28)(29)(30)(31)(32)(33)(34)(35)(36)(37)(38)(39) peptide indicated that nasal immunization with the combined HBs and HBc formulation activates antigen specific CD8 + cells, consistent with a strong Th1 response. This result is in line with the finding that B cells that bind viral capsids can prime cytotoxic T cells.…”

“…After washing, the cells were counted and distributed at 2 × 10 6 cells/mL in 10 mL RPMI FCS in 25 cm 2 flasks (Nalge-Nunc, Rochester, NY, USA) and stimulated with 10 mg/mL of peptide S (28)(29)(30)(31)(32)(33)(34)(35)(36)(37)(38)(39) . After culturing for 4 days in 5% CO 2 , half of the total medium was substituted and new medium containing 2 × 10 4 U/mL of IL-2 was added.…”

“…The cells were extensively washed with medium, resuspended in RPMI-1640 (Gibco, Scotland, UK) supplemented with 10% FCS, 2 mmol/L glutamine, 2 mmol/L piruvate, 50 mmol/L 2-mercaptoethanol and antibiotics (complete medium) and counted. Meanwhile, H-2 d mastocytome cells p815 were pulsed for 1 h at 37 ° C, 5% CO 2 in complete medium with 10 mmol/L S (28)(29)(30)(31)(32)(33)(34)(35)(36)(37)(38)(39) peptide IPQSLDSWWTSL from HBsAg. 33 After incubation p815 cells were further incubated for another 15 min with Mitomycin C (Sigma).…”

“…immunization with HBsAg in alum. The IFN-γ ELISPOT assay was conducted using the immunodominant HBs (28)(29)(30)(31)(32)(33)(34)(35)(36)(37)(38)(39) CTL peptide from HBsAg as previously described. Measurement of lymphoproliferative activity revealed the induction of strong proliferative responses in systemic compartments against both HBsAg and HBcAg after nasal-combined immunization.…”

Methazolamide Calcium Phosphate Nanoparticles in an Ocular Delivery System

“…Mixing was accomplished by slowly adding 7.5 ml of 12.5 mM calcium chloride solution and 7.5 ml of 12.5 mM dibasic sodium phosphate solution into 1.5 ml of 15.6 mM sodium citrate solution at 4°C by continuous stirring (2000 rpm) for 48 h. The mixture was sonicated (frequency 22 kHz; power 120 W) for 30 min, the dispersion wasˆltered and was kept at -4°C for further use. 16,17) Preparation of Methazolamide CaP Nanoparticles…”