Immunostimulatory properties of a novel adjuvant administered with inactivated influenza virus vaccine.

The immunopotentiating activity of a new delivery system was investigated comparatively to Alhydrogel adjuvant, as an antiviral inactivated vaccine after one injection. The efficiency of the new formulation (BioMed) was evaluated with an inactivated porcine strain of influenza (A/Sw/IN/1726/88 H1N1) in the pig model. The first assessment criteria was the follow-up of selected immunological parameters such as, antibody levels, lymphoproliferation, double positive CD4+CD8+ T lymphocytes and cytokine production (IL-2, IL-4, IFN-gamma). The second criteria was the estimate of the protection level of animals exposed to a homologous challenge of 50 PID50 one month after a single immunizing or control injection. In the BioMed group of animals, 4 pigs (out of 6) were free of macroscopic lesion, while lesions could be seen in all individuals of other groups and virus was isolated in only one animal, whereas all other animals of other groups had virus in their lungs. This better protection of BioMed animals seems to be correlated mainly with higher levels of antibodies and to a lesser extent with a slightly better CMI response and probably with the production of memory CD4+CD8+ T cells.

Synthesis and characterization of new permanently charged poly(amidoammonium) salts and evaluation of their DNA complexes for gene transport.

A new series of linear and permanently charged poly(amidoammonium) salts were synthesized in order to investigate the influence of their ionic and hydrophobic contents on both the cytotoxicity and the transfection mediated by polycation-DNA complexes. The poly(amidoammonium) salts were prepared by chemical modification of a parent poly(amidoamine) containing two tertiary amino groups per structural unit: one incorporated into the main chain and the other fixed at the end of a short bismethylene spacer. The permanent charges were introduced through a quaternization reaction involving iodomethane or 1-iodododecane as an alkylating agent. Under appropriate conditions, the methylation reaction was found to be regioselective, allowing the quaternization of either the side chains or both the side chains and the backbone. Under physiological salt conditions (150 mM NaCl), all of the poly(amidoammonium) salts self-assembled with DNA to form complexes. High proportions of highly quaternized polycation provided better defined morphology to the polycation-DNA complexes. Complexes formed from unquaternized polycation were less cytotoxic than branched poly(ethyleneimine) (25 kDa). At high polycation-DNA weight ratios, the introduction of permanent charges generated a significant increase in the cytotoxicity, but no patent correlation could be established with the amount and the position of the permanent charges. Only complexes formed from polycations with quaternized backbone were able to generate significant gene expression, which was putatively attributed to a better defined toroidal-like morphology together with a higher stability, as suggested by zeta potential measurements. The incorporation of dodecane side chains on highly charged polycations severely amplified the cytotoxicity so that, in return, the transfection level was dramatically affected.

Vaccination of cattle with a DNA plasmid encoding the bovine viral diarrhoea virus major glycoprotein E2.

Bovine viral diarrhoea virus (BVDV) is an economically important pathogen of cattle that is ubiquitously distributed worldwide. In this study, cattle were immunized by intramuscular injections with plasmid DNA expressing the BVDV type 1 major glycoprotein E2. Animals either received injections of naked DNA (N-DNA) or DNA in cationic liposomes (L-DNA). Both DNA preparations induced virus-specific neutralizing antibodies in vaccinates, although the response was much lower in N-DNA-immunized animals. N-DNA-vaccinated animals also showed virus-specific lymphocyte proliferation responses to type 1, live BVDV in vitro, whereas L-DNA vaccination induced no such responses. After 16 weeks, DNA-vaccinated and mock-vaccinated animals were challenged with a USDA-certified BVDV type 1 strain. Four significant observations were made: (1) N-DNA-vaccinated calves showed limited protection from virus challenge, (2) L-DNA-vaccinated animals did not show any signs of protection, (3) the challenge induced strong memory responses in the production of serum neutralizing antibodies to both genotypes (type 1 and 2 of BVDV), and (4) the challenge induced a mucosal memory response in nasal secretions of both L- and N-DNA-vaccinated animals.