Physicochemical and immunological characterization of N,N,N-trimethyl chitosan-coated whole inactivated influenza virus vaccine for intranasal administration.
OBJECTIVE
The purpose of this study was the development and physicochemical and immunological characterization of intranasal (i.n.) vaccine formulations of whole inactivated influenza virus (WIV) coated with N,N,N-trimethyl chitosan (TMC).


METHODS
Synthesized TMCs with a degree of quarternization of 15% (TMC15) or 37% (TMC37) were tested in vitro for their ability to decrease the transepithelial resistance (TEER) of an epithelial cell monolayer. TMC15- and TMC37-coated WIV (TMC15-WIV and TMC37-WIV) were characterized by zeta potential measurements, dynamic light scattering, electron microscopy and gel permeation chromatography. Mice were vaccinated i.n. with selected vaccine formulations and immunogenicity was determined by measuring serum hemagglutination inhibition (HI) and serum IgG, IgG1 and IgG2a/c titers. Also a pulse-chase study with TMCs in solution administered i.n. 2 h prior to WIV was performed. Protective efficacy of vaccination was determined by an aerosol virus challenge.


RESULTS
TMC37 induced a reversible decrease in TEER, suggesting the opening of tight junctions, whereas TMC15 did not affect TEER. Simple mixing of (negatively charged) WIV with TMC15 or TMC37 resulted in positively charged particles with TMCs being partially bound. Intranasal immunization with TMC37-WIV or TMC15-WIV induced stronger HI, IgG, IgG1 and IgG2a/c titers than WIV alone. TMC37-WIV induced the highest immune responses. Both TMC15-WIV and TMC37-WIV provided protection against challenge, whereas WIV alone was not protective. Intranasal administration of TMC prior to WIV did not result in significant immune responses, indicating that the immunostimulatory effect of TMC is primarily based on improved i.n. delivery of WIV.


CONCLUSIONS
Coating of WIV with TMC is a simple procedure to improve the delivery and immunogenicity of i.n. administered WIV and may enable effective i.n. vaccination against influenza.
Hagenaars
Niels
N
Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, The Netherlands.
Mastrobattista
Enrico
E
Verheul
Rolf J
RJ
Mooren
Imke
I
Glansbeek
Harrie L
HL
Heldens
Jacco G M
JG
van den Bosch
Han
H
Jiskoot
Wim
W
D000281:Administration, Intranasal; D000818:Animals; D018938:Caco-2 Cells; D002470:Cell Survival / Q000187:drug effects; D048271:Chitosan / Q000737:chemistry* / Q000276:immunology* / Q000633:toxicity; D017097:Electric Impedance; D005260:Female; D006801:Humans; D007114:Immunization; D007252:Influenza Vaccines / Q000008:administration & dosage* / Q000737:chemistry / Q000276:immunology*; D029521:Alphainfluenzavirus / Q000276:immunology*; D051379:Mice; D008810:Mice, Inbred C57BL; D010316:Particle Size
D007252:Influenza Vaccines; C118071:N-trimethyl chitosan chloride; D048271:Chitosan