Conventional vaccine strategies have been highly efficacious for several decades in reducing mortality and morbidity due to infectious diseases. The bane of conventional vaccines, such as those that include whole organisms or large proteins, appear to be the inclusion of unnecessary antigenic load that, not only contributes little to the protective immune response, but complicates the situation by inducing allergenic and/or reactogenic responses. Peptide vaccines are an attractive alternative strategy that relies on usage of short peptide fragments to engineer the induction of highly targeted immune responses, consequently avoiding allergenic and/or reactogenic sequences. Conversely, peptide vaccines used in isolation are often weakly immunogenic and require particulate carriers for delivery and adjuvanting. In this article, we discuss the specific advantages and considerations in targeted induction of immune responses by peptide vaccines and progresses in the development of such vaccines against various diseases. Additionally, we also discuss the development of particulate carrier strategies and the inherent challenges with regard to safety when combining such technologies with peptide vaccines.

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Chlamydia has been shown to evade host-specific IFN-γ-mediated bacterial killing; however, IFN-γ-deficient mice exhibit suboptimal late phase vaginal Chlamydia muridarum clearance, greater dissemination, and oviduct pathology. These findings introduce constraints in understanding results from murine chlamydial vaccination studies in context of potential implications to humans. In this study, we used mice deficient in either IFN-γ or the IFN-γ receptor for intranasal vaccination with a defined secreted chlamydial Ag, chlamydial protease-like activity factor (CPAF), plus CpG and examined the role of IFN-γ derived from adoptively transferred Ag-specific CD4+ T cells in protective immunity against genital C. muridarum infection. We found that early Ag-specific IFN-γ induction and CD4+ T cell infiltration correlates with the onset of genital chlamydial clearance. Adoptively transferred IFN-γ competent CPAF-specific CD4+ T cells failed to enhance the resolution of genital chlamydial infection within recipient IFN-γ receptor-deficient mice. Conversely, IFN-γ production from adoptively transferred CPAF-specific CD4+ T cells was sufficient in IFN-γ-deficient mice to induce early resolution of infection and reduction of subsequent pathology. These results provide the first direct evidence that enhanced anti-C. muridarum protective immunity induced by Ag-specific CD4+ T cells is dependent upon IFN-γ signaling and that such cells produce sufficient IFN-γ to mediate the protective effects. Additionally, MHC class II pathway was sufficient for induction of robust protective anti-C. muridarum immunity. Thus, targeting soluble candidate Ags via MHC class II to CD4+ T cells may be a viable vaccine strategy to induce optimal IFN-γ production for effective protective immunity against human genital chlamydial infection.

Genital chlamydial infections lead to severe upper reproductive tract pathology in a subset of untreated women. We demonstrated previously that TNF-α producing CD8+ T cells contribute significantly to chlamydial upper genital tract pathology in female mice. Additionally, we observed minimal chlamydial oviduct pathology develops in OT-1 transgenic (OT-1) mice, wherein CD8+ T cell repertoire is restricted to recognition of the ovalbumin peptide Ova257–264, suggesting that non-Chlamydia-specific CD8+ T cells may not be responsible for chlamydial pathogenesis. In the current study, we evaluated whether antigen-specific CD8+ T cells mediate chlamydial pathology. Groups of wild type C57BL/6J (WT), OT-1 mice, and OT-1 mice replete with WT CD8+ T cells (1×106 cells/mouse intravenously) were infected intravaginally with C. muridarum (5 × 104 IFU/mouse). Serum total anti-Chlamydia antibody and total splenic anti-Chlamydia IFN-γ and TNF-α responses were comparable among the three groups of animals. However, Chlamydia-specific IFN-γ and TNF-α production from purified splenic CD8+ T cells of OT-1 mice was minimal, whereas responses in OT-1 mice replete with WT CD8+ T cells were comparable to those in WT animals. Vaginal chlamydial clearance was comparable between the three groups of mice. Importantly, the incidence and severity of oviduct and uterine horn pathology was significantly reduced in OT-1 mice but reverted to WT levels in OT-1 mice replete with WT CD8+ T cells. Collectively, these results demonstrate that Chlamydia-specific CD8+ T cells contribute significantly to upper genital tract pathology.

An important consideration for antichlamydial vaccine development is the induction of cross-serovar protection, since multiple serovars (D to L) of Chlamydia trachomatis cause genital infections. We have shown previously that vaccination with C. trachomatis-derived recombinant chlamydial protease-like activity factor (rCPAF) induced significant earlier resolution of Chlamydia muridarum infection and reduced oviduct pathology. However, the vaccinated mice continued to shed chlamydiae for up to 2 weeks after challenge. In this study, C. trachomatis serovar D recombinant proteins, such as recombinant major outer membrane protein (rMOMP), recombinant inclusion membrane protein A (rIncA), and rCPAF were administered intranasally, individually or in combinations, with murine interleukin-12 (IL-12) as an adjuvant, and cross-species immunity against intravaginal C. muridarum infection was examined. Immunization with rCPAF plus IL-12 (rCPAF؉IL-12), compared to immunization with rIncA؉IL-12 or rMOMP؉IL-12, induced the greatest antigen-specific gamma interferon production from purified CD4 ؉ T cells and concurrently enhanced serum antibody production. All (100%) the animals vaccinated with rCPAF؉IL-12 alone or in any combination completely resolved the infection by day 18 after challenge compared to animals vaccinated with rIncA؉IL-12 (50%), rMOMP؉IL-12 (33%), or phosphate-buffered saline (mock vaccinated; 0%). Moreover, oviduct pathology in mice vaccinated by any regimen that included rCPAF, but not rMOMP؉IL-12 or rIncA؉IL-12 alone, was markedly reduced compared to mock-immunized animals. The addition of rMOMP and/or rIncA did not significantly enhance the rCPAF؉IL-12-induced effect on bacterial clearance or oviduct pathology. These results suggest a greater conservation of protective linear antigenic epitopes within CPAF than MOMP or IncA across the examined serovars and the need to identify other highly conserved antigens for use with rCPAF in a multisubunit recombinant vaccine.

Mice deficient in B-cells (μmT mice) were used to evaluate the role of antibody in enhanced chlamydial clearance and reduction of pathology afforded by vaccination with recombinant chlamydial protease-like activity factor (rCPAF). Enhanced, but comparable, chlamydial clearance was observed in μmT and wild type (WT) mice after rCPAF+CpG vaccination. Chlamydia-induced pathology was present in mock-immunized animals, but at significantly greater levels in μmT than WT mice, whereas vaccinated μmT and WT mice exhibited similar reductions in pathology. Thus, antibodies may play a role in protection against chlamydial pathology after primary infection, but were largely dispensable in rCPAF+CpG induced chlamydial clearance and reduction in pathology.

We previously have shown the efficacy of recombinant (r) chlamydial protease-like activity factor (CPAF) vaccination against hydrosalpinx development following primary genital chlamydial challenge. In this study, we evaluated further the protection induced by rCPAF vaccination against infertility. Following primary challenge, fertility levels were not significantly different between the mock- and CPAF-vaccinated and Chlamydia alone challenged mice. However, following secondary genital chlamydial challenge, mock (PBS) immunized mice displayed a significant reduction of fertility compared to age-matched naïve mice, while mice vaccinated intranasally with rCPAF+CpG displayed significant prevention of infertility. These results suggest that hydrosalpinx may be a reliable indicator of impending infertility, and that rCPAF is a promising candidate to prevent infertility resulting from repeated genital chlamydial infections.

Collectively, these results demonstrate that TNFR2-bearing CD8(+) T cells and TNFR1-bearing non-CD8(+) T cells contribute significantly to oviduct pathology following genital chlamydial infection.