Louisa Tao

Protective mucosal immunity elicited by targeted iliac lymph node immunization with a subunit SIV envelope and core vaccine in macaques.

Prevention of sexually transmitted HIV infection was investigated in macaques by immunization with a recombinant SIV (simian immunodeficiency virus) envelope gp120 and core p27 vaccine. In two independent series of experiments, we used the novel targeted iliac lymph node (TILN) route of immunization, aiming close to the iliac lymph nodes draining the genitorectal mucosa. Rectal challenge with the SIVmac 32H J5 molecular clone in two series induced total protection in four out of seven macaques immunized by TILN, compared with infection in 13 of 14 unimmunized macaques or immunized by other routes (P = 0.025). The remaining three macaques showed either a decrease in viral load (>90%) or transient viremia, indicating that all seven TILN–immunized macaques showed total or partial protection (P = 0.001). Protection was associated with significant increase in the iliac lymph nodes of lgA antibody–secreting cells to p27 (P < 0.02), CD8–suppressor factor (P< 0.01), and the chemokines RANTES and MIP–1β (P< 0.01).

Immunogenicity of the Extracellular Domains of C-C Chemokine Receptor 5 and the In Vitro Effects on Simian Immunodeficiency Virus or HIV Infectivity

The C-C chemokine receptor CCR5 serves an important function in chemotaxis of lymphocytes, monocytes, and dendritic cells. CCR5 is also the major coreceptor in most macrophage-tropic HIV-1 infections. Immunization of rhesus macaques with a baculovirus-generated CCR5 construct or peptides derived from the sequences of the four extracellular domains of CCR5 elicited IgG and IgA Abs, inhibition of SIV replication, and CD4+ T cell proliferative responses to three of the extracellular domains of CCR5. The immune sera reacted with cell surface CCR5 expressed on HEK 293 cells. T and B cell epitope mapping revealed major and minor T and B cell epitopes in the N-terminal, first, and second loops of CCR5. The three C-C chemokines, RANTES, macrophage-inflammatory protein-1α, and macrophage-inflammatory protein-1β, were up-regulated by immunization with the CCR5-derived peptides, and the cell surface expression of CCR5 was decreased. The CCR5 Abs were complementary to the C-C chemokines in inhibiting HIV replication in vitro. Immunization with the four extracellular domains of CCR5 suggests that three of them are immunogenic, with maximal T cell responses being elicited by the second loop peptide. However, maximal Abs to the cell surface CCR5 or viral inhibitory Abs in vitro were induced by the N-terminal peptide. Up-regulation of the three C-C chemokines and down-modulation of cell surface CCR5 were elicited by the second loop, N-terminal, and first loop peptides. The data suggest that a dual mechanism of C-C chemokines and specific Abs may engage and down-modulate the CCR5 coreceptors and prevent in vitro HIV or SIV replication.

Generation of CD8+ T cell-generated suppressor factor and beta-chemokines by targeted iliac lymph node immunization in rhesus monkeys challenged with SHIV-89.6P by the rectal route.

The targeted lymph node (TLN) immunization strategy was investigated in macaques, in order to determine the efficacy in generating secretory, systemic, and cellular immune responses, CD8+ T cell-generated suppressor factors, and beta-chemokines. TLN immunization of the rectal and genital mucosa-associated iliac lymph nodes (TILNs) was compared with axillary TLN immunization (TAxLN) using HIV-1 MN/LAI gp140env and SIV p27gag in alum. Significantly higher immune responses, as well as CD8+ T cell-generated anti-SIV factors and the beta-chemokines RANTES, MIP-1alpha, and MIP-1beta, were elicited by iliac as compared with axillary TLN immunization. The immune responses induced by TLN immunization were examined for their capacity to prevent rectal mucosal infection by the pathogenic dual-tropic SHIV-89.6P. Despite significant secretory, serum, cellular, and beta-chemokine responses, the macaques were infected by SHIV-89.6P. Whether the lack of protection was associated with the antigenic unrelatedness of SHIV-89.6P to the immunizing HIV-1 MN/LAI gp140 or to the virus utilizing CXCR4 to a much greater extent than CCR5, remains to be determined.

The common mucosal immune system for the reproductive tract: basic principles applied toward an AIDS vaccine

Abstract The concept of the Collaborative Mucosal Immunization Research Group for AIDS (CMIG) was originally conceived by the AIDS Vaccine Branch, National Institute of Allergy and Infectious Diseases (NIAID) in order to provide support for a cooperative research environment for the development of mucosal immunity to AIDS. We have purposely organized five groups of investigators at five different locations to determine how effective mucosal immunity to AIDS can be optimally approached. CMIG recognizes that both rectal (homosexual) as well as vaginal (heterosexual) infections with HIV are two of the major ways that AIDS currently disseminates through the human population. Thus, we have chosen the SIV model of infection of rhesus macaques, but more importantly the CMIG have joined two of our five components in order to use the significant expertise developed for mucosal transmission of SIV and immunity. Thus, we have brought the extensive expertise with vaginal and rectal immunization and immunity to spread [Drs. Chris Miller and Marta Marthas, California Regional Primate Research Center (CRPRC), Davis and Drs. Thomas Lehner and Martin Cranage, United Medical and Dental School Guys Hospital, London and the Centre for Applied Microbiology and Research (Guys/CAMR)]. Two additional components were added in order to perform mucosal immune response studies required to develop and to optimize a mucosal vaccine. First, extensive CD4+ T helper (Th) cell (e.g., Th1 and Th2) and CD8+ T cell subset studies are a major effort of the coordinating group at the University of Alabama at Birmingham (Drs. Hiroshi Kiyono and Jerry R. McGhee). This component is closely interacting with both the CRPRC and Guys/CAMR components in terms of SIV-specific CD4+ and CD8+ T cell subset responses. For example, SIV-specific CTL responses are jointly examined using different techniques by CRPRC, Guys/CAMR and UAB investigators. Further, it is also important to examine a balance between SIV-specific and Th1 and Th2 cell responses following mucosal immunization since the Th cell-derived cytokines are essential for the induction of appropriate antigen-specific mucosal immune responses. This issue is currently being extensively examined by the CMIG effort and a summary of our findings is discussed in this review. A major question in mucosal immunity involves the functions of secretory IgA (S-IgA) antibodies and this area is of particular importance in rectal and reproductive tract immunity. A novel and exciting in vitro epithelial cell assay system is used to study how effectively S-IgA neutralizes SIV infection (Drs. John Huang, John Nedrud and Michael Lamm, Case Western Reserve, Cleveland). A clear advantage of this CMIG effort is the unique expertise in design of mucosal delivery systems for an AIDS vaccine. We are using state-of-the-art recombinant bacteria, i.e.. rSalmonella and rVibrios for mucosal immunization [Drs. Yichen Lu and Bryan Roberts, Virus Research Institute (VRI), Boston]. In addition, we are also testing other mucosal delivery systems including DNA vaccine, microspheres, cholera toxin (CT) and CT-B, recombinant poliovirus, and immune complexes. These studies represent the first efforts to induce not only Th cell mediated S-IgA responses, but also CTL responses to SIV in primates immunized with different mucosal vector delivery systems. In order to focus our effort for the induction of SIV-specific immune responses following mucosal immunization, investigators from the CMIG are attempting to understand the induction and regulation of antigen-specific immune responses in rhesus macaques mucosally immunized with different preparations of SIV vaccines.

Generation of diversity in the hierarchy of T-cell epitope responses following different routes of immunization with simian immunodeficiency virus protein.

ObjectivesTo examine whether the route of immunization determines the hierarchy of T-cell epitope proliferative responses in macaques. DesignMacaques were immunized with a recombinant simian immunodeficiency virus (SIV) p27 core protein by the intramuscular, male and female genital or rectal route, each of which was augmented by oral immunization, and by the novel targeted lymph-node immunization route. Overlapping peptides were used to identify the proliferative T-cell epitopes and to determine their hierarchy in the circulation, spleen and lymph nodes. MethodsT-cell epitope mapping of the proliferative responses was studied in short-term cell lines. Dendritic cells and macrophages were enriched by metrizamide gradient and adherence to plastic, respectively. ResultsIntramuscular immunization elicited in the circulating T cells a hierarchy of T-cell epitopes within four peptides in the following descending order of frequency: peptides 121–140 (57.9%), 41–60 (28.9%), 61–80 (18.9%) and 101–120 (5.4%). The hierarchy of these four T-cell epitope responses differed significantly with each of the five routes of immunization, when circulating (P<0.001), splenic (P<0.02-<0.001) or iliac lymph-node cells (P<0.001) were analysed. The effect of antigen-presenting cells was then investigated and enriched dendritic cells were more effective than macrophages in processing and presenting the p27 antigen and the immunodominant (121–140) and 61–80 T-cell epitopes. ConclusionsThe route of immunization may determine the hierarchy of T-cell epitopes in the lymph nodes draining the mucosa in the circulating and splenic lymphocytes. The diversity of T-cell epitopes may affect the control of HIV at different anatomical sites, the administration route of the vaccine, and selection of polypeptides or recombinant antigens for immunization.

CD8-suppressor factor and β-chemokine function as a complementary mechanism to cognate immunity

Protection against SIV or HIV infection requires specific antibodies and T-cell immune responses. However, a complementary mechanism may be involved, in which CD8-suppressor factors (CD8-SF) and the constitutive beta-chemokines may prevent the virus binding and replicating. Indeed, there is evidence that targeted iliac lymph node (TILN) immunisation with SIVgp120 and p27 or xenoimmunisation with SIV grown in human T-cells generates CD8-SF, RANTES, MIP-1alpha and MIP-1beta which are significantly correlated with protection from SIV infection by the rectal mucosal or intravenous route, respectively. Inhibition of SIV replication in vitro is dependent on the concentration of beta-chemokines generated by immunisation. The critical level for inhibition of SIV replication appears to be higher for rectal mucosal than intravenous challenge by SIV. The mechanism of protection in vivo has not been elucidated but it is likely that the beta-chemokines bind to CCR5 coreceptors which are internalised. Thus, CCR5 coreceptors are either blocked or not expressed on the cell surface for SIV to bind and infect.