Invasive amoebiasis due to Entamoeba histolytica is a leading cause of death and morbidity worldwide. Immunity to intestinal infection would prevent the occurrence of amebic colitis and amebic liver abscess (ALA); currently, no vaccine exists. A 260 kDa galactose-inhabitable lectin mediates the binding of E. histolytica trophozoites to colonic mucins and epithelial cells; the lectin consists of 170kDa and 35kDa subunits. The 170kDa subunit contains the galactose-binding activity and is highly antigenic and immunogenic. The genes encoding the 170kDa subunit have been cloned and sequenced for E. histolytica and the non-pathogenic species, E. dispar. A 52kDa cysteine-rich galactose-binding site and is highly antigenic. Subcutaneous vaccination with LC3 provides immunity in the gerbil model of ALA, oral immunization with LC3 and cholera toxin induces are adherence/inhibitory anti-lectin mucosal secretory IgA response in BALB/c mice.
The objectives of this proposal are to identify LC3 IgA antibody and T-cell epitopes associated with immunity to E. histolytica intestinal infection and invasive amoebiasis and to develop an efficacious LC3-derived amoebiasis subunit vaccine.
The specific aims and methods are: <ol>
<LI>To define the LC3 epitopes associated with acquired immunity to intestinal infection and invasive amoebiasis by utilizing purified LC3 fragment sin ELISA with serum, fecal and salivary IgA antibodies from immune individuals by defining IgA epitopes in D. dispar lectin, by mapping LC3 T-cell epitopes using PBMC from immune subjects, and by identifying LC3 epitopes recognized by baboon IgA following E. histolytica infection; and
<LI>To develop an LC3 subunit vaccine that elicits an adherence-inhibitory intestinal secretory IgA response and/or a protective cellular immune response by establishing a dose response to oral vaccination in baboons with LC3 and cholera holotoxin for induction of anti-LC3 IgA antibodies, by defining the efficacy in baboons with LC3 and cholera holotoxin for induction of anti-LC3 IgA antibodies, by defining the efficacy in baboons of LC3 vaccination against oral challenge with E. histolytica cysts, by determining antigen-specific cellular immune responses in baboons; and lastly, by identifying LC3 fragments or derived peptides that are sufficient to serve as an efficacious subunit vaccine in the primate model. Successful completion of thee studies is dependent upon collaboration with investigators in Project 2, use of the Hybridoma Core, and ongoing interactions with the "Mucosal Immunity and Infection Project" faculty.
The proposed studies will greatly enhance our understanding of human mucosal immunity to E. histolytica, leading to development of an effective amoebiasis subunit vaccine.