Protection against Intestinal Amebiasis by a Recombinant Vaccine Is Transferable by T Cells and Mediated by Gamma Interferon

We have previously shown that vaccination with purified Entamoeba histolytica Gal/GalNAc lectin or recombinant subunits can protect mice from intestinal amebiasis upon intracecal challenge. In this study, we demonstrated with adoptive-transfer experiments that this lectin vaccine protection is mediated by T cells but not serum. The cell-mediated immune (CMI) response was characterized by significant gamma interferon (IFN-γ), interleukin 12 (IL-12), IL-2, IL-10, and IL-17 production. To move toward a human vaccine, we switched to a recombinant protein and tested a range of adjuvants and routes appropriate for humans. We found that subcutaneous delivery of LecA with IDRI''s adjuvant system EM014 elicited a potent Th1-type CMI profile and provided significant protection, as measured by culture negativity (79% efficacy); intranasal immunization with cholera toxin provided 56% efficacy; and alum induced a Th2-type response that protected 62 to 68% of mice. Several antibody and CMI cytokine responses were examined for correlates of protection, and prechallenge IFN-γ⁺ or IFN-γ-, IL-2-, and tumor necrosis factor alpha-triple-positive CD4 cells in blood were statistically associated with protection. To test the role of IFN-γ in LecA-mediated protection, we neutralized IFN-γ in LecA-immunized mice and found that it abrogated the protection conferred by vaccination. These data demonstrate that CMI is sufficient for vaccine protection from intestinal amebiasis and reveal an important role for IFN-γ, even in the setting of alum.The enteric protozoan parasite Entamoeba histolytica is the causative pathogen of amebic dysentery and liver abscess that affects millions of people worldwide. Bangladeshi children experience a 40% annual incidence of E. histolytica infection (24), and evidence of prior E. histolytica infection can be detected in 8.4% of the general population in Mexico (6). Despite the availability of effective antibiotics, the World Health Organization estimates that up to 100,000 deaths occur annually, highlighting the need for alternate approaches to control amebiasis. One approach is to develop a vaccine to prevent intestinal infection (26).Several vaccine candidates for amebiasis have been proposed (48), including the serine-rich E. histolytica protein, peroxiredoxin, the EhCP112 molecule, and the galactose/N-acetyl-d-galactosamine-inhibitable lectin (Gal/GalNAc lectin). A significant body of work has focused on the latter: vaccination with either parasite-purified Gal/GalNAc lectin (10, 29, 32, 38, 40) or recombinant lectin subunits has provided protection in rodent models against amebic liver abscess and amebic colitis (29, 37, 46, 47, 53). Although these results are encouraging, two limitations remain. First, in most of these vaccine studies, the adjuvants and delivery routes are not compatible with eventual use in humans. Second, the mechanisms of amebiasis vaccine-mediated protection are still not fully understood. For instance, in the intestinal model, there was an association between the presence of an antiparasite lectin fecal immunoglobulin A (IgA) response and subsequent protection, but the association did not extend to the recombinant antigen, and fecal IgA-negative mice remained statistically protected, suggesting that other immune mechanisms exist (29). Indeed, there is increasing evidence for a role of cell-mediated immunity (CMI) in protection from intestinal amebiasis (14, 20, 28). We have found that gamma interferon (IFN-γ), the canonical Th1 cytokine, can clear intestinal amebic infection in CBA mice (21). In this study, we demonstrate for the first time that CMI plays a critical role in lectin-elicited protective immunity to intestinal amebic infection. A variety of vaccine adjuvants and delivery routes were tested for their effectiveness in protection and in eliciting CMI, and the tests demonstrated a clear role for CMI and IFN-γ in lectin-based vaccine protection.