2 months of disulfiram. cART was discontinued 8 weeks after last vaccination. IFN-c ELISPOT was used to assess immuno- genicity and proviral reservoir was ...
Conclusions: This study shows that Tregs represent an important fraction of HIV-specific T cells. Tregs impact the frequency and the capability of effector cells to control viral replication. Thus the ability to measure the inducibility of Tregs and/or to modulate those responses prior to vaccination is important for more efficient strategies in future vaccines.
P24.02 Seminal Plasma Modulates Dendritic Cell Function Favoring the Generation of CD251/FOXP31 T-cells Antonela Merlotti, Maria Julia Ruiz, Fernando Erra Dı´az, Ezequiel Dantas, Augusto Varese, Gabriel Duette, Pehuen Pereyra, Ernst Glenda, Federico Remes Lenicov, Jorge Geffner, Juan Sabatte´ Instituto de Investigaciones Biome´dicas en Retrovirus y SIDA, Universidad de Buenos Aires/CONICET, Capital Federal, Argentina Background: Unprotected sexual intercourse is the most common mode of HIV-1 transmission being semen the most important vector for this infection. Dendritic cells (DCs) are abundantly located on mucosal surfaces and play different roles during HIV infection: promote HIV spread by boosting CD4 + T cell infection and activate the HIV specific adaptive immune response. As semen has been shown to promote immune tolerance in different models, we hypothesize that components present in plasma seminal (SP) might modulate DC function promoting a tolerogenic immune response. To test this, we study the ability of complete SP to modulate DC function and the ability of these cells to induce CD25 + /FOXP3 + regulatory T cells. Methods: SP was obtained from healthy donors. Monocyte derived DCs were cultured during 24 hs with SP samples (diluted 1/ 100) in the absence or presence of LPS (10ng/ml). DC phenotype was studied by flow cytometry. Cytokine secretion was measured in culture supernatants by ELISA. After SP treatment, DCs were cultured with allogeneic CD4 + T cells and the induction of CD25 + /FOXP3 + T cells was analyzed by flow cytometry. Results: We found that SP inhibited IL-12, IL-6, TNF-alpha and IL-1, but not IL-10 production by LPS stimulated DCs (percent of inhibition respect to LPS alone: 81.2 + / -11.07% p < 0.0001 for IL-12, 43.45 + / -2.87% p < 0.001 for IL-6, 69.8 + / -17.31% p < 0.001 for TNF-alpha, 37.5 + / -13.15% p < 0.05 for IL-1, 6.1 + / -15.6% p = 0.6 for IL-10). SP also boosted the ability of LPS stimulated DCs to induce CD25 + /FOXP3 + T cells (PS + LPS = 20,4% vs LPS alone = 6%, p < 0,05). We observed no changes on the expression of HLA-DR, CD80, CD86, CD83, CD40 and CD1a on immature or LPS-maturated DCs after incubation with SP. Conclusions: SP modulates DC function, inhibiting the secretion of pro-inflammatory cytokines and favoring the induction of CD25 + /FOXP3 + T cells. In this way, we speculate that SP might modulate the adaptive immune response against sexual transmitted pathogens.
P24.03 In Vivo Viral Control in a HLA-B*35:01 Homozygous Individual after the Vaccine-induced Response to a Welldefined, HIV Gag-derived HLA-B*35 CTL Epitope Miriam Rosas1, Beatriz Mothe1,2, Nu´ria Climent3, Maria C Puertas1, Javier Martinez-Picado1,4,5, Felipe Garcia3, Christian Brander1,4,5, and the RISVAC03 Trial Investigator Team
1
IrsiCaixa AIDS Research Institute - HIVACAT, Badalona, Spain, University of Vic and Central Catalonia, Barcelona, Spain, 3 Hospital Clinic-HIVACAT, IDIBAPS, Barcelona, Spain, 4Institucio´ Catalana de Recerca i Estudis Avanc¸ats (ICREA), Barcelona, Spain, 5University of Vic and Central Catalonia, Vic, Spain 2
Background: Virus-specific CD8 T cell responses to epitopes restricted by HLA-B*35:01 are generally believed to be ineffective in mediating in vivo control of HIV infection. We report a case of a patient homozygous for HLA-B*35:01 who showed successful viral control after vaccination with MVA-B vaccine combined with a drug to reactivate HIV-1 replication (disulfiram), and who subsequently underwent an analytical treatment interruption (ATI). Methods: Therapeutic vaccination consisted of 3 intramuscular injections of MVA-B at 0,4,16 weeks and a 4th dose followed by 2 months of disulfiram. cART was discontinued 8 weeks after last vaccination. IFN-c ELISPOT was used to assess immunogenicity and proviral reservoir was determined over time. Viral rebound dynamics were assessed during ATI. Results: The patient had a past history of high viral load set point (362,000 copies/ml) and cART was initiated during chronic infection. After ATI the patient remained with low level viral load < 200 copies/ml for > 24 weeks without showing a significant decay in CD4 T-cell counts. At baseline, the patient showed a broad (19 different specificities) and strong (9,189 SFC/106PBMC) T cell response, which increased to 14,470 SFC/106PBMC after three vaccinations. Responses to three novel T-cell epitopes present in the vaccine insert (Nef A*03QK10, Gag A*03-RK9 and Pol B*35-VY10) were induced upon 3 vaccination. A dominant HLA-B*35:01 restricted response to the Gag-p24 B*35-PY9 epitope (PPIPVGDIY) of 3,330 SFC/ 106PBMC was detected before ATI. No changes in proviral reservoir or viral expression (mRNA) were observed in CD4+ T-cells after disulfiram treatment. Conclusions: The expansion of a dominant response towards the HLA-B*35- restricted Gag RY9 epitope could potentially explain the observed viral control on this subject suggesting that certain HLA-B*35:01 restricted responses may have the potential to significantly contribute to viral control, providing important guidance for vaccine immunogen design covering non-beneficial HLA alleles.
P24.04 Evolution of Polyfunctional and Proliferative CD81 T-cell Responses from Early to Chronic HIV-1 Infection Meika EI Richmond1,2, Sandra A. Kiazyk1,2, Lyle R. Mckinnon3, Charles Wachihi4, Makubo Kimani4, Joshua Kimani4, Francis A. Plummer1,4,5, T. Blake Ball1,2,4 1 University of Manitoba, Medical Microbiology, Winnipeg, MB, Canada, 2Public Health Agency of Canada, National Lab for HIV Immunology, Winnipeg, MB, Canada, 3University of KwaZulu-Natal, Centre for the Programme of AIDS Research in South Africa, Durban, Kenya, 4University of Nairobi, Nairobi, Kenya, 5Public Health Agency of Canada, National Microbiology Lab, Winnipeg, MB, Canada
Background: The limited success of HIV vaccine candidates to date highlights our need to better characterize protective cellmediated immunity (CMI). HIV-infected subjects that experience slower progression to AIDS, provide a valuable model for the study of CMI responses that may be capable of controlling
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HIV. Previous work has demonstrated that these individuals maintain stronger HIV-specific CD8 + T cell proliferation and polyfunctionality compared to progressing controls. However it is unclear whether these CD8 + T cell characteristics are responsible better disease outcomes or if they are merely a consequence of the individuals high CD4 + T cells and low viral loads as a result of protection by other means. Methods: Here we assessed the evolution of CD8 + polyfunctional and proliferative responses from early to chronic HIV-1 infection in 26 HIV infected individuals following seroconversion using multiparameter flow cytometry. We hypothesized that CD8 + T cells will become increasingly polyfunctional and proliferative from early to chronic infection and patients that mount and maintain an early polyfunctional and proliferative response will have a better disease outcome. Results: Our data suggests that the polyfunctional and proliferative capacity of CD8 + T cells from early to chronic infection follows a distinct pattern, and further that CD8 + T cell responses vary substantially between individuals in the chronic phase of infection. Case study analysis of individuals CD8 + T responses over time suggests that individuals with moderate polyfunctionality in the early phase of infection go on to maintain healthy CD4 + T cell counts. Conclusions: Our data lends support to the hypothesis that polyfunctional and proliferative CD8 + T cells are the cause of slow HIV disease progression. Identification of whether polyfunctional responses and strong proliferative capacity is the cause or consequence of HIV control will be needed for the comprehensive evaluation of HIV vaccine candidates.
P24.05 Identification of CD81 T-cell Epitopes that Associate with Distinct Functionality, Proliferation and Polyfunctionality Meika EI Richmond1,2, Sandra A. Kiazyk1,2, Lyle R. Mckinnon3, Billy Nyanga4, Charles Wachihi4, Makubo Kimani4, Joshua Kimani2,4, Francis A. Plummer2,4,5, T. Blake Ball1,2,4 1
Public Health Agency of Canada, National Lab for HIV Immunology, Winnipeg, MB, Canada, 2University of Manitoba, Medical Microbiology, Winnipeg, MB, Canada, 3University of KwaZulu-Natal, Centre for the Programme of AIDS Research in South Africa, Durban, Kenya, 4University of Nairobi, Nairobi, Kenya, 5Public Health Agency of Canada, National Microbiology Lab, Winnipeg, MB, Canada Background: Understanding correlates CD8 + T cell protection against HIV infection and progressive disease is essential for informing effective vaccine development, design and evaluation. CD8 + T cell responses with a robust polyfunctional and proliferative component are strongly linked to better disease outcomes. However, the specificity of polyfunctional and proliferative CD8 + T cell responses has not been thoroughly investigated. This study provides a better understanding of the fine specificity of HIV-specific CD8 + T cell responses. Methods: Here we have selected eleven HIV-1 Clade A p24 epitopes of interest, which were previously identified during our comprehensive fine epitope mapping study, for further characterization. Responses to these epitopes were measured in 83 chronically HIV-infected individuals from a Kenyan female sex worker cohort, using multiple cytokines/chemokines and proliferation. Results: Responses to eleven epitopes were consistent with our previous findings, and most epitope-specific responses were
IFN-g negative (64%) while many responses were polyfunctional (38%). We identified epitopes that preferentially elicited specific cytokines/chemokines (p < 0.001) while other epitopes trend towards eliciting a proliferative response. Additionally our data suggests that particular epitopes are associated with polyfunctionality (p < 0.0001). Notably, we identified an epitope (EEKAFSPEV) that associates with MIP-1b, polyfunctionality and proliferation, all attributes linked with slower disease progression. Conclusions: Importantly, we show that, at a cohort level, particular epitopes preferentially elicit specific qualities of CD8 + T cell responses in preference to others. This suggests there is potential to identify specific epitopes that elicit protective polyfunctional and/or proliferative CD8 + T cell responses. Such ‘protective’ epitopes could be incorporated into a vaccine to express distinct CD8 + T cell epitopes and induce a more effective CD8 + T cell response.
P24.06 HIV-Specific CD81 T-cell Expansion Potential, but Not Cytotoxic Capacity, Is Associated with Reduced Set Point Viral Loads in Ad5/HIV Vaccinees Stephen A. Migueles1, Nicole Frahm2, Sushila A. Toulmin1, Elizabeth P. Kelly1, Bennett A. Peterson1, Sarah A. Johnson1, M J. McElrath2, Mark Connors1 1 NIAID, NIH, Bethesda, MD, United States, 2Fred Hutchinson Cancer Research Center, Seattle, WA, United States
Background: HIV-specific CD8 + T-cell cytotoxicity is a robust correlate of control in chronic infection, but low cytotoxicity was induced by Ad5/HIV vaccines. Although some vaccinees maintain control of HIV, the ability of expansion potential or cytotoxic capacity to predict this control has not been examined in the NIAID HVTN 502 Step Study. Methods: Pre-infection PBMCs of 37 vaccinees were analyzed in a blinded manner. In a novel real-time imaging-based assay, cytotoxic responses were measured as the elimination of HIVGFP-infected CD4 + T-cell targets by 6-day re-stimulated CD8 + T cells. Responses were also measured in LTNP/EC (n = 19) and progressors (n = 20). True E/T ratios were derived from measures of IFN-gamma + CD107a + CD8 + T cells and HIV GFP + targets. Responses were correlated with set point HIV RNA levels. Results: Cytotoxicity by confocal imaging was highly correlated with flow cytometric-based assays (r = 0.8, p < 0.001). Cytotoxic responses of Step vaccinees were significantly lower than those of LTNP/EC (27.9% v. 68.3%, respectively; p < 0.001) and progressors (43.9%, p < 0.001). Low cytotoxic capacity was not simply due to lower frequency as the per-cell cytotoxic capacity of Step subjects was significantly lower than that of LTNP/EC (p < 0.01). Although the two subjects with HIV RNA set points < 400 copies/mL had high responses, overall cytotoxic capacity did not correlate with viral loads (r = -0.24, p > 0.05). However, expansion potential, estimated by Day 6 IFN-gamma + CD107a + cell frequencies, was weakly correlated with HIV RNA levels (r = -0.42, p = 0.01). Conclusions: Pre-infection HIV-specific CD8 + T-cell cytotoxic capacity of Ad5/HIV vaccine recipients in Step was low and did not predict set point HIV RNA levels. A better understanding of the reasons that Ad5/HIV induces such low levels of per-cell cytotoxic capacity may provide critical insights for induction of an efficacious CD8 + T-cell response by HIV/AIDS vaccines.
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