Preparation and characterization of innovative protein-coated poly(methylmethacrylate) core-shell nanoparticles for vaccine purposes.
OBJECTIVE
This study aims at developing novel core-shell poly(methylmethacrylate) (PMMA) nanoparticles as a delivery system for protein vaccine candidates.


METHODS
Anionic nanoparticles consisting of a core of PMMA and a shell deriving from Eudragit L100/55 were prepared by an innovative synthetic method based on emulsion polymerization. The formed nanoparticles were characterized for size, surface charge and ability to reversibly bind two basic model proteins (Lysozyme, Trypsin) and a vaccine relevant antigen (HIV-1 Tat), by means of cell-free studies. Their in vitro toxicity and capability to preserve the biological activity of the HIV-1 Tat protein were studied in cell culture systems. Finally, their safety and immunogenicity were investigated in the mouse model.


RESULTS
The nanoparticles had smooth surface, spherical shape and uniform size distribution with a mean diameter of 220 nm. The shell is characterized by covalently bound carboxyl groups negatively charged at physiological pH, able to reversibly adsorb large amounts (up to 20% w/w) of basic proteins (Lysozyme, Trypsin and HIV-1 Tat), mainly through specific electrostatic interactions. The nanoparticles were stable, not toxic to the cells, protected the HIV-1 Tat protein from oxidation, thus preserving its biological activity and increasing its shelf-life, and efficiently delivered and released it intracellularly. In vivo experiments showed that they are well tolerated and elicit strong immune responses against the delivered antigen in mice.


CONCLUSIONS
This study demonstrates that these new nanoparticles provide a versatile platform for protein surface adsorption and a promising delivery system particularly when the maintenance of the biologically active conformation is required for vaccine efficacy.
Voltan
Rebecca
R
Department of Histology, Microbiology and Medical Biotechnology, Section of Microbiology, University of Padova, Via A. Gabelli 63, 35122, Padova, Italy.
Castaldello
Arianna
A
Brocca-Cofano
Egidio
E
Altavilla
Giuseppe
G
Caputo
Antonella
A
Laus
Michele
M
Sparnacci
Katia
K
Ensoli
Barbara
B
Spaccasassi
Silvia
S
Ballestri
Marco
M
Tondelli
Luisa
L
D016915:AIDS Vaccines / Q000737:chemistry / Q000276:immunology / Q000378:metabolism / Q000633:toxicity; D000180:Acrylic Resins / Q000737:chemistry* / Q000633:toxicity; D000818:Animals; D000917:Antibody Formation; D002470:Cell Survival / Q000187:drug effects; D002626:Chemistry, Pharmaceutical; D004305:Dose-Response Relationship, Drug; D004337:Drug Carriers; D004339:Drug Compounding; D004355:Drug Stability; D005260:Female; D006367:HeLa Cells; D006801:Humans; D007111:Immunity, Cellular; D051379:Mice; D008810:Mice, Inbred C57BL; D009113:Muramidase / Q000737:chemistry; D053758:Nanoparticles; D010084:Oxidation-Reduction; D010316:Particle Size; D019904:Polymethyl Methacrylate / Q000737:chemistry* / Q000633:toxicity; D011485:Protein Binding; D011489:Protein Denaturation; D012995:Solubility; D013997:Time Factors; D014357:Trypsin / Q000737:chemistry; D054322:tat Gene Products, Human Immunodeficiency Virus / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism / Q000633:toxicity
D016915:AIDS Vaccines; D000180:Acrylic Resins; D004337:Drug Carriers; C446821:Eudragit L100-55; C421254:human immunodeficiency virus type 1 Tat vaccine; D054322:tat Gene Products, Human Immunodeficiency Virus; D019904:Polymethyl Methacrylate; D009113:Muramidase; D014357:Trypsin