The objective of this application is to evaluate the mucosal adjuvant activities of the Escherichia coli Type II enterotoxins, LT-IIa and LT-IIb. Experiments in the laboratory of the applicant demonstrated that LT-IIa and LT-IIb induce different and distinctive patterns of enhanced immune responses, and that those patterns are profoundly different from those induced by cholera toxin (CT). For example, whereas CT used as an adjuvant induces predominantly a T helper 2-type response based on antibody isotype and cytokine patterns, Type II enterotoxins, particularly LT-IIb, induce both T helper 1 and T helper 2 responses. These data provide strong evidence that LT-IIa, LT-IIb, and CT induce their adjuvant activities using different cellular and molecular mechanisms. As such, the Type II toxins provide an elegant set of tools for investigating the mechanisms of mucosal adjuvant induction. Although related in structure, LT-IIa, LT-IIb and CT bind to different sets of cell surface receptors. It is hypothesized that the distinctive adjuvant activities of the toxins are governed by their receptor-binding specificities. To test this hypothesis, the adjuvant activities of the Type II toxins will be analyzed in a mucosal mouse model using AgI/II of the oral pathogen Streptococcus mutans as a model antigen. Both antibody and cellular responses will be assessed. These studies will be facilitated by a collection of receptor-binding mutants, hybrid molecules, and chimeric toxins that are available in this laboratory. Immunization studies will be combined with immunohistological investigations of lymphoid tissue to begin to investigate the cellular component of toxin-induced adjuvant activity. Confocal microscopy will be used to identify the immunocompetent cells in the nasal lymphoid tissue and the draining lymph nodes that initially interact with the toxins after intranasal inoculation. As a further means to correlate adjuvant induction with toxin/cell interactions, immunocompetent cells taken from nasal lymphoid tissue will be classified for expression of toxin-specific surface receptors using flow cytometry analysis. Finally, the potential of non-toxic chimeric Type II proteins as adjuvant/antigen delivery vehicles will be evaluated. At the conclusion of these studies, the laboratory will be well positioned to evaluate the therapeutic potential of the Type II toxins as mucosal adjuvants in the subsequent production of new vaccines that will protect against pathogens that infect the oral, gastric and urogenital mucosae.
Showing the most recent 10 out of 31 publications
