Novel mucosal DNA-MVA HIV vaccination in which DNA-IL-12 plus Cholera Toxin B subunit (CTB) cooperates to enhance cellular systemic and mucosal genital tract immunity

Induction of local antiviral immune responses at the mucosal portal surfaces where HIV-1 and other viral pathogens are usually first encountered remains a primary goal for most vaccines against mucosally acquired viral infections. Exploring mucosal immunization regimes in order to find optimal vector combinations and also appropriate mucosal adjuvants in the HIV vaccine development is decisive. In this study we analyzed the interaction of DNA-IL-12 and cholera toxin B subunit (CTB) after their mucosal administration in DNA prime/MVA boost intranasal regimes, defining the cooperation of both adjuvants to enhance immune responses against the HIV-1 Env antigen. Our results demonstrated that nasal mucosal DNA/MVA immunization schemes can be effectively improved by the co-delivery of DNA-IL-12 plus CTB inducing elevated HIV-specific CD8 responses in spleen and more importantly in genital tract and genito-rectal draining lymph nodes. Remarkably, these CTL responses were of superior quality showing higher avidity, polyfunctionality and a broader cytokine profile. After IL-12+CTB co-delivery, the cellular responses induced showed an enhanced breadth recognizing with higher efficiency Env peptides from different subtypes. Even more, an in vivo CTL cytolytic assay demonstrated the higher specific CD8 T-cell performance after the IL-12+CTB immunization showing in an indirect manner its potential protective capacity. Improvements observed were maintained during the memory phase where we found higher proportions of specific central memory and T memory stem-like cells T-cell subpopulations. Together, our data show that DNA-IL-12 plus CTB can be effectively employed acting as mucosal adjuvants during DNA prime/MVA boost intranasal vaccinations, enhancing magnitude and quality of HIV-specific systemic and mucosal immune responses.Fil: Maeto, Cynthia Alejandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Rodríguez, Ana María. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Holgado, María Pía. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Falivene, Juliana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Gherardi, Maria Magdalena. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; Argentin

Development of vaccines for HIV-1: relevance of subtype-specific cellular immunity

Han pasado casi 30 años de la detección de los primeros casos de infección con HIV-1 y aún no se ha conseguido desarrollar una vacuna efectiva y segura. A pesar del impacto positivo sobre la pandemia que se ha conseguido gracias a los avances en la terapia antirretroviral (TARV), el HIV/sida sigue constituyendo un grave problema para la salud pública, especialmente en los países en desarrollo, donde es difícil el acceso al tratamiento. En el mundo, 33 millones de personas viven con el virus del sida, mientras que en la Argentina se calcula que habría unos 120 000 infectados. Uno de los desafíos para lograr una vacuna contra el HIV es la variabilidad viral. El grupo M, responsable de la pandemia, se encuentra dividido en 10 subtipos y varios sub-subtipos, además de las 48 formas recombinantes circulantes y más de cien formas recombinantes únicas. La epidemia de HIV en nuestro país es tan compleja como en el resto del mundo, con la co-circulación principalmente de virus pertenecientes al subtipo B y recombinantes BF (CRF12_BF y derivadas). A pesar de la cantidad de trabajos dedicados a la caracterización de la respuesta inmune y al desarrollo de vacunas, no queda claro cuál es el impacto de la variabilidad en la elección del antígeno. Trabajos realizados en nuestro laboratorio demuestran el papel que juega la inmunidad celular con respecto a las variantes recombinantes BF, tanto en humanos como en modelos animales. Estos resultados son de importancia en el desarrollo de futuras vacunas para nuestra región.It has been almost 30 years since the detection of the first HIV-1 cases and yet an effective and safe vaccine has not been developed. Although, advances in antiretroviral therapy (HAART) have produced a major impact on the pandemic, and even though HIV/aids remains a major concern for developing countries, where access to therapy is limited. The last report from UNAIDS notified 33 million people living with HIV/aids, worldwide, while in Argentina it is estimated that 120 000 persons have been infected. One of the challenges to address and ultimately overcome when developing a vaccine is the high variability of HIV-1. The M group, responsible for the pandemic, is divided into 10 subtypes and several sub-subtypes, in addition to the 48 circulating recombinant forms (CRF) and over one hundred unique recombinant forms (URF). The HIV epidemic in Argentina is as complex as in the rest of the world, characterized by the high prevalence of infections caused by subtype B and BF variants. Despite the wide range of publications focused on the immune response against HIV as well as to vaccine development, how to overcome variability on vaccine antigen selection is still unclear. Studies performed in our laboratory showed the impact of the immunogenicity of BF recombinant variants, both in humans and in animal models. These results are of great concern in vaccine development for our region.Fil: Rodríguez, Ana María. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Microbiología. Centro Nacional de Referencia del Sida; ArgentinaFil: Turk, Gabriela Julia Ana. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Microbiología. Centro Nacional de Referencia del Sida; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Pascutti, María Fernanda. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Microbiología. Centro Nacional de Referencia del Sida; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Falivene, Juliana. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Microbiología. Centro Nacional de Referencia del Sida; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Gherardi, Maria Magdalena. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Microbiología. Centro Nacional de Referencia del Sida; Argentin

Th17 and Th17/Treg ratio at early HIV infection associate with protective HIV-specific CD8+ T-cell responses and disease progression

The aim of this study was to analyze Th17 and Treg subsets and their correlation with anti-HIV T-cell responses and clinical parameters during (acute/early) primary HIV infection (PHI) and up to one year post-infection (p.i). Samples from 14 healthy donors (HDs), 40 PHI patients, 17 Chronics, and 13 Elite controllers (ECs) were studied. The percentages of Th17 and Treg subsets were severely altered in Chronics, whereas all HIV-infected individuals (including ECs) showed Th17/Treg imbalance compared to HDs, in concordance with higher frequencies of activated CD8+ T-cells (HLA-DR+/CD38+). Better clinical status (higher CD4 counts, lower viral loads and activation) was associated with higher Th17 and lower Treg levels. We found positive correlations between Th17 at baseline and anti-HIV CD8+ T-cell functionality: viral inhibitory activity (VIA) and key polyfunctions (IFN-γ+/CD107A/B+) at both early and later times p.i, highlighting the prognostic value of Th17 cells to preserve an effective HIV T-cell immunity. Th17/Treg ratio and the IL-17 relative mean fluorescence intensity (rMFI of IL-17) were also positively correlated with VIA. Taken together, our results suggested a potential link between Th17 and Th17/Treg ratio with key HIV-specific CD8+ T-cell responses against the infection.Fil: Falivene, Juliana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Ghiglione, Yanina Alexandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Laufer, Natalia Lorna. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; Argentina. Gobierno de la Ciudad de Buenos Aires. Hospital General de Agudos "Juan A. Fernández"; ArgentinaFil: Socías, María Eugenia. Fundación Huésped; ArgentinaFil: Holgado, María Pía. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Ruiz, Maria Julia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Maeto, Cynthia Alejandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Figueroa, María Inés. Fundación Huésped; ArgentinaFil: Giavedoni, Luis D.. Texas Biomedical Research Institute; Estados UnidosFil: Cahn, Pedro. Gobierno de la Ciudad de Buenos Aires. Hospital General de Agudos "Juan A. Fernández"; Argentina. Fundación Huésped; ArgentinaFil: Salomon, Horacio Eduardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Sued, Omar Gustavo. Fundación Huésped; ArgentinaFil: Turk, Gabriela Julia Ana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Gherardi, Maria Magdalena. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; Argentin

Early Skewed Distribution of Total and HIV-Specific CD8+ T-Cell Memory Phenotypes during Primary HIV Infection Is Related to Reduced Antiviral Activity and Faster Disease Progression

The important role of the CD8+ T-cells on HIV control is well established. However, correlates of immune protection remain elusive. Although the importance of CD8+ T-cell specificity and functionality in virus control has been underscored, further unraveling the link between CD8+ T-cell differentiation and viral control is needed. Here, an immunophenotypic analysis (in terms of memory markers and Programmed cell death 1 (PD-1) expression) of the CD8+ T-cell subset found in primary HIV infection (PHI) was performed. The aim was to seek for associations with functional properties of the CD8+ T-cell subsets, viral control and subsequent disease progression. Also, results were compared with samples from Chronics and Elite Controllers. It was found that normal maturation of total and HIV-specific CD8+ T-cells into memory subsets is skewed in PHI, but not at the dramatic level observed in Chronics. Within the HIV-specific compartment, this alteration was evidenced by an accumulation of effector memory CD8+ T (TEM) cells over fully differentiated terminal effector CD8+ T (TTE) cells. Furthermore, higher proportions of total and HIV-specific CD8+ TEM cells and higher HIV-specific TEM/(TEM+TTE) ratio correlated with markers of faster progression. Analysis of PD-1 expression on total and HIV-specific CD8+ T-cells from PHI subjects revealed not only an association with disease progression but also with skewed memory CD8+ T-cell differentiation. Most notably, significant direct correlations were obtained between the functional capacity of CD8+ T-cells to inhibit viral replication in vitro with higher proportions of fully-differentiated HIV-specific CD8+ TTE cells, both at baseline and at 12 months post-infection. Thus, a relationship between preservation of CD8+ T-cell differentiation pathway and cell functionality was established. This report presents evidence concerning the link among CD8+ T-cell function, phenotype and virus control, hence supporting the instauration of early interventions to prevent irreversible immune damage.Fil: Ghiglione, Yanina Alexandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Falivene, Juliana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Ruiz, Maria Juliz. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Laufer, Natalia Lorna. Gobierno de la Ciudad de Buenos Aires. Hospital General de Agudos ; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Socías, María Eugenia. Fundación Huésped; Argentina. Gobierno de la Ciudad de Buenos Aires. Hospital General de Agudos ; ArgentinaFil: Cahn, Pedro. Gobierno de la Ciudad de Buenos Aires. Hospital General de Agudos ; Argentina. Fundación Huésped; ArgentinaFil: Giavedoni, Luis. Southwest National Primate Research Center; Estados UnidosFil: Sued, Omar Gustavo. Fundación Huésped; Argentina. Gobierno de la Ciudad de Buenos Aires. Hospital General de Agudos ; ArgentinaFil: Gherardi, Maria Magdalena. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Salomon, Horacio Eduardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Turk, Gabriela Julia Ana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; Argentin

IL-12 DNA Displays efficient adjuvant effects improving immunogenicity of Ag85A in DNA prime/MVA boost immunizations

Mycobacterium tuberculosis (Mtb) infection is one of the leading causes of death worldwide. The Modified Vaccinia Ankara (MVA) vaccine vector expressing the mycobacterial antigen 85A (MVA85A) was demonstrated to be safe, although it did not improve BCG efficacy, denoting the need to search for improved tuberculosis vaccines. In this work, we investigated the effect of IL-12 DNA -as an adjuvant- on an Ag85A DNA prime/MVA85A boost vaccination regimen. We evaluated the immune response profile elicited in mice and the protection conferred against intratracheal Mtb H37Rv challenge. We observed that the immunization scheme including DNA-A85A+DNA-IL-12/MVA85A induced a strong IFN-γ production to Ag85A in vitro, with a significant expansion of IFN-γ+CD4+ and IFN-γ+CD8+ anti-Ag85A lymphocytes. Furthermore, we also detected a significant increase in the proportion of specific CD8+CD107+ T cells against Ag85A. Additionally, inclusion of IL-12 DNA in the DNA-A85A/MVA85A vaccine scheme induced a marked augment in anti-Ag85A IgG levels. Interestingly, after 30 days of infection with Mtb H37Rv, DNA-A85A+DNA-IL-12/MVA85A vaccinated mice displayed a significant reduction in lung bacterial burden. Together, our findings suggest that IL-12 DNA might be useful as a molecular adjuvant in an Ag85A DNA/MVA prime-boost vaccine against Mtb infection.Instituto de BiotecnologíaFil: Morelli, María Paula. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina.Fil: Del Medico Zajac, Maria Paula. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular; Argentina.Fil: Pellegrini, Joaquín Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina.Fil: Amiano, Nicolás Oscar. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina.Fil: Tateosian, Nancy Liliana. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina.Fil: Calamante, Gabriela. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular; Argentina.Fil: Gherardi, Maria Magdalena. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; Argentina.Fil: García, Verónica Edith. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina.Fil: Pellegrini, Joaquín Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina.Fil: Pellegrini, Joaquín Miguel. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica; Argentina.Fil: Pellegrini, Joaquín Miguel. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica; ArgentinaFil: Morelli, María Paula. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica; Argentina.Fil: Morelli, María Paula. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica; ArgentinaFil: Del Medico Zajac, Maria Paula. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Amiano, Nicolás Oscar. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica; Argentina.Fil: Amiano, Nicolás Oscar. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica; ArgentinaFil: Tateosian, Nancy Liliana. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica; Argentina.Fil: Tateosian, Nancy Liliana. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica; ArgentinaFil: Calamante, Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Gherardi, Maria Magdalena. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: García, Verónica Edith. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica; Argentina.Fil: García, Verónica Edith. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica; Argentin

Phenotype, polyfunctionality, and antiviral activity of in vitro stimulated CD8+ T-Cells From HIV+ subjects who initiated cART at different time-points after acute infection

Since anti-HIV treatment cannot cure the infection, many strategies have been proposed to eradicate the viral reservoir, which still remains as a major challenge. The success of some of these strategies will rely on the ability of HIV-specific CD8+ T-cells (CD8TC) to clear reactivated infected cells. Here, we aimed to investigate the phenotype and function of in vitro expanded CD8TC obtained from HIV+ subjects on combination antiretroviral therapy (cART), either initiated earlier (median = 3 months postinfection, ET: Early treatment) or later (median = 20 months postinfection, DT: Delayed treatment) after infection. Peripheral blood mononuclear cells from 12 DT and 13 ET subjects were obtained and stimulated with Nef and Gag peptide pools plus IL-2 for 14 days. ELISPOT was performed pre- and post-expansion. CD8TC memory/effector phenotype, PD-1 expression, polyfunctionality (CD107a/b, IFN-γ, IL-2, CCL4 (MIP-1β), and/or TNF-α production) and antiviral activity were evaluated post-expansion. Magnitude of ELISPOT responses increased after expansion by 103 times, in both groups. Expanded cells were highly polyfunctional, regardless of time of cART initiation. The memory/effector phenotype distribution was sharply skewed toward an effector phenotype after expansion in both groups although ET subjects showed significantly higher proportions of stem-cell and central memory CD8TCs. PD-1 expression was clustered in HIV-specific effector memory CD8TCs, subset that also showed the highest proportion of cytokine-producing cells. Moreover, PD-1 expression directly correlated with CD8TC functionality. Expanded CD8TCs from DT and ET subjects were highly capable of mediating antiviral activity, measured by two different assays. Antiviral function directly correlated with the proportion of fully differentiated effector cells (viral inhibition assay) as well as with CD8TC polyfunctionality and PD-1 expression (VITAL assay). In sum, we show that, despite being dampened in subjects on cART, the HIV-specific CD8TC response could be selectively stimulated and expanded in vitro, presenting a high proportion of cells able to carry-out multiple effector functions. Timing of cART initiation had an impact on the memory/effector differentiation phenotype, most likely reflecting how different periods of antigen persistence affected immune function. Overall, these results have important implications for the design and evaluation of strategies aimed at modulating CD8TCs to achieve the HIV functional cure.Fil: Salido, Jimena Patricia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Ruiz, María Julia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Trifone, César Ariel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Figueroa, María Inés. Fundación Huésped; ArgentinaFil: Caruso, Maria Paula. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Gherardi, Maria Magdalena. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Sued, Omar Gustavo. Fundación Huésped; ArgentinaFil: Salomon, Horacio Eduardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Laufer, Natalia Lorna. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Ghiglione, Yanina Alexandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Turk, Gabriela Julia Ana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; Argentin

Phenotype, Polyfunctionality, and Antiviral Activity of in vitro Stimulated CD8+ T-Cells From HIV+ Subjects Who Initiated cART at Different Time-Points After Acute Infection

Since anti-HIV treatment cannot cure the infection, many strategies have been proposed to eradicate the viral reservoir, which still remains as a major challenge. The success of some of these strategies will rely on the ability of HIV-specific CD8+ T-cells (CD8TC) to clear reactivated infected cells. Here, we aimed to investigate the phenotype and function of in vitro expanded CD8TC obtained from HIV+ subjects on combination antiretroviral therapy (cART), either initiated earlier (median = 3 months postinfection, ET: Early treatment) or later (median = 20 months postinfection, DT: Delayed treatment) after infection. Peripheral blood mononuclear cells from 12 DT and 13 ET subjects were obtained and stimulated with Nef and Gag peptide pools plus IL-2 for 14 days. ELISPOT was performed pre- and post-expansion. CD8TC memory/effector phenotype, PD-1 expression, polyfunctionality (CD107a/b, IFN-γ, IL-2, CCL4 (MIP-1β), and/or TNF-α production) and antiviral activity were evaluated post-expansion. Magnitude of ELISPOT responses increased after expansion by 103 times, in both groups. Expanded cells were highly polyfunctional, regardless of time of cART initiation. The memory/effector phenotype distribution was sharply skewed toward an effector phenotype after expansion in both groups although ET subjects showed significantly higher proportions of stem-cell and central memory CD8TCs. PD-1 expression was clustered in HIV-specific effector memory CD8TCs, subset that also showed the highest proportion of cytokine–producing cells. Moreover, PD-1 expression directly correlated with CD8TC functionality. Expanded CD8TCs from DT and ET subjects were highly capable of mediating antiviral activity, measured by two different assays. Antiviral function directly correlated with the proportion of fully differentiated effector cells (viral inhibition assay) as well as with CD8TC polyfunctionality and PD-1 expression (VITAL assay). In sum, we show that, despite being dampened in subjects on cART, the HIV-specific CD8TC response could be selectively stimulated and expanded in vitro, presenting a high proportion of cells able to carry-out multiple effector functions. Timing of cART initiation had an impact on the memory/effector differentiation phenotype, most likely reflecting how different periods of antigen persistence affected immune function. Overall, these results have important implications for the design and evaluation of strategies aimed at modulating CD8TCs to achieve the HIV functional cure

Early Gag Immunodominance of the HIV-Specific T-Cell Response during Acute/Early Infection Is Associated with Higher CD8 T-Cell Antiviral Activity and Correlates with Preservation of the CD4 TCell Compartment

The important role of the CD8 T-cell response on HIV control is well established. Moreover, the acute phase of infection represents a proper scenario to delineate the antiviral cellular functions that best correlate with control. Here, multiple functional aspects (specificity, ex vivo viral inhibitory activity [VIA] and polyfunctionality) of the HIV-specific CD8 T-cell subset arising early after infection, and their association with disease progression markers, were examined. Blood samples from 44 subjects recruited within 6 months from infection (primary HIV infection [PHI] group), 16 chronically infected subjects, 11 elite controllers (EC), and 10 healthy donors were obtained. Results indicated that, although Nef dominated the anti-HIV response during acute/early infection, a higher proportion of early anti-Gag T cells correlated with delayed progression. Polyfunctional HIV-specific CD8 T cells were detected at early time points but did not associate with virus control. Conversely, higher CD4 T-cell set points were observed in PHI subjects with higher HIV-specific CD8 T-cell VIA at baseline. Importantly, VIA levels correlated with the magnitude of the anti-Gag cellular response. The advantage of Gag-specific cells may result from their enhanced ability to mediate lysis of infected cells (evidenced by a higher capacity to degranulate and to mediate VIA) and to simultaneously produce IFN-. Finally, Gag immunodominance was associated with elevated plasma levels of interleukin 2 (IL-2) and macrophage inflammatory protein 1 (MIP-1). All together, this study underscores the importance of CD8 T-cell specificity in the improved control of disease progression, which was related to the capacity of Gag-specific cells to mediate both lytic and nonlytic antiviral mechanisms at early time points postinfection.Fil: Turk, Gabriela Julia Ana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomedicas en Retrovirus y Sida; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Microbiologia. Centro Nacional de Referencia del Sida; ArgentinaFil: Ghiglione, Yanina Alexandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomedicas en Retrovirus y Sida; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Microbiologia. Centro Nacional de Referencia del Sida; ArgentinaFil: Falivene, Juliana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomedicas en Retrovirus y Sida; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Microbiologia. Centro Nacional de Referencia del Sida; ArgentinaFil: Socias, María Eugenia. Fundación Huésped; ArgentinaFil: Laufer, Natalia Lorna. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomedicas en Retrovirus y Sida; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Microbiologia. Centro Nacional de Referencia del Sida; Argentina. Gobierno de la Ciudad de Buenos Aires. Hospital General de Agudos;Fil: Coloccini, Romina Soledad. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomedicas en Retrovirus y Sida; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Microbiologia. Centro Nacional de Referencia del Sida; ArgentinaFil: Rodríguez, Ana María. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomedicas en Retrovirus y Sida; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral;Fil: Ruiz, María Julia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomedicas en Retrovirus y Sida; ArgentinaFil: Pando, Maria de Los Angeles. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomedicas en Retrovirus y Sida; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Microbiologia. Centro Nacional de Referencia del Sida; ArgentinaFil: Giavedoni, Luis David. Texas Biomedical Research Institute. Southwest National Primate Research Center. Department of Virology and Immunology; Estados UnidosFil: Cahn, Pedro. Fundación Huésped; Argentina. Gobierno de la Ciudad de Buenos Aires. Hospital General de Agudos;Fil: Sued, Omar Gustavo. Fundación Huésped; ArgentinaFil: Salomon, Horacio Eduardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomedicas en Retrovirus y Sida; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Microbiología; ArgentinaFil: Gherardi, Maria Magdalena. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomedicas en Retrovirus y Sida; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Microbiologia. Centro Nacional de Referencia del Sida; Argentin

IL-12 and GM-CSF in DNA/MVA Immunizations against HIV-1 CRF12_BF Nef Induced T-Cell Responses With an Enhanced Magnitude, Breadth and Quality

In Argentina, the HIV epidemic is characterized by the co-circulation of subtype B and BF recombinant viral variants. Nef is an HIV protein highly variable among subtypes, making it a good tool to study the impact of HIV variability in the vaccine design setting. We have previously reported a specific cellular response against NefBF with low cross-reactivity to NefB in mice. The aim of this work was to analyze whether the co-administration of IL-12 and GM-CSF, using DNA and MVA vaccine vectors, could improve the final cellular response induced. Mice received three DNA priming doses of a plasmid that express NefBF plus DNAs expressing IL-12 and/or GM-CSF. Afterwards, all the groups were boosted with a MVAnefBF dose. The highest increase in the magnitude of the NefBF response, compared to that induced in the control was found in the IL-12 group. Importantly, a response with higher breadth was detected in groups which received IL-12 or GM-CSF, evidenced as an increased frequency of recognition of homologous (BF) and heterologous (B) Nef peptides, as well as a higher number of other Nef peptide pools representing different viral subtypes. However, these improvements were lost when both DNA cytokines were simultaneously administered, as the response was focused against the immunodominant peptide with a detrimental response towards subdominant epitopes. The pattern of cytokines secreted and the specific-T-cell proliferative capacity were improved in IL-12 and IL-12+GM-CSF groups. Importantly IL-12 generated a significant higher T-cell avidity against a B heterologous peptide