Development of nasal vaccines based on cholera toxin B subunit conjugated to bacterial polysaccharides

Abstract

Objective: Cholera toxin B subunit (CTB) is a strong mucosal immunogen and has also been used as a mucosal carrier for other protein antigens, but not previously for polysaccharide antigens. The aim of this thesis was to see if CTB could be used as a mucosal carrier for polysaccharides, particularly for nasal vaccinations. As CTB has not been used for intranasal vaccinations before, volunteers were vaccinated with different doses of CTB as a nasal spray. Dextran and Haemophilus influenzae type b capsular polysaccharide (HibCPS) were coupled to CTB and used for immunization studies in mice.Methods: Volunteers were vaccinated with 10, 100 and 1000 µg of CTB as a nasal spray. Side effects were recorded and the antibody responses in serum and nasal and vaginal secretions were measured in ELISA. Dextran and HibCPS were conjugated to CTB and the conjugates were used for mucosal immunizations of mice. The antibody responses in mice were measured in serum and in extracts of organs.Results: The 10 and 100 µg doses did not give any side effects in the volunteers, while the 1000 µg dose gave local side effects. The 100 µg dose elicited IgA and IgG antibody responses in serum and nasal and vaginal secretions. Dextran was coupled to CTB. Intranasal and peroral immunizations with the conjugate elicited local and systemic antibody responses to dextran, in contrast to the immunizations with unconjugated dextran, which did not elicit any antibodies. Pre-existing local immunity to CTB suppressed the antibody response to dextran after immunization with conjugate. HibCPS could not be efficiently conjugated to CTB directly. As an alternative, HibCPS was first coupled to tetanus toxoid and then to CTB. The HibTTCTB conjugate elicited local and systemic antibody responses after intranasal immunization, but the HibTT conjugate did not elicit any local antibodies after intranasal immunization. Conclusions: It is possible to obtain a local and systemic antibody response to a bacterial polysaccharide after intranasal immunization if it is coupled to CTB. CTB can be given safely to humans intranasally in an immunogenic dose. Thus, bacterial polysaccharides conjugated to CTB can potentially be used as nasal vaccines.