Monica Montes

Dendritic cell-based therapeutic vaccine elicits polyfunctional HIV-specific T-cell immunity associated with control of viral load.

Efforts aimed at restoring robust immune responses limiting human immunodeficiency virus (HIV)‐1 replication therapeutically are warranted. We report that vaccination with dendritic cells generated ex vivo and loaded with HIV lipopeptides in patients (n = 19) on antiretroviral therapy was well tolerated and immunogenic. Vaccination increased: (i) the breadth of the immune response from 1 (1–3) to 4 (2–5) peptide‐pool responses/patient (p = 0.009); (ii) the frequency of functional T cells (producing at least two cytokines among IFN‐γ, TNF‐α, and IL‐2) from 0.026 to 0.32% (p = 0.002) and from 0.26 to 0.35% (p = 0.005) for CD4+ and CD8+ T cells, respectively; and (iii) the breadth of cytokines secreted by PBMCs upon antigen exposure, including IL‐2, IFN‐γ, IL‐21, IL‐17, and IL‐13. Fifty percent of patients experienced a maximum of viral load (VL) 1 log10 lower than the other half following antiretroviral treatment interruption. An inverse correlation was found between the maximum of VL and the frequency of polyfunctional CD4+ T cells (p = 0.007), production of IL‐2 (p = 0.006), IFN‐γ (p = 0.01), IL‐21 (p = 0.006), and IL‐13 (p = 0.001). These results suggest an association between vaccine responses and a better control of viral replication. These findings will help in the development of strategies for a functional cure for HIV infection.

Development of a HIV-1 lipopeptide antigen pulsed therapeutic dendritic cell vaccine

In the search for a therapeutic HIV-1 vaccine, we describe herein the development of a monocyte-derived dendritic cell (DC) vaccine loaded with a mixture of HIV-1-antigen lipopeptides (ANRS HIV-LIPO-5 Vaccine). LIPO-5 is comprised of five HIV-1-antigen peptides (Gag(17-35), Gag(253-284), Nef(66-97), Nef(116-145), and Pol(325-355)), each covalently linked to a palmitoyl-lysylamide moiety. Monocytes enriched from HIV-1-infected highly active antiretroviral therapy (HAART)-treated patients were cultured for three days with granulocyte-macrophage colony-stimulating factor and alpha-interferon. At day 2, the DCs were loaded with ANRS HIV-LIPO-5 vaccine, activated with lipopolysaccharide, harvested at day 3 and frozen. Flow cytometry analysis of thawed DC vaccines showed expression of DC differentiation markers: CD1b/c, CD14, HLA-DR, CD11c, co-stimulatory molecule CD80 and DC maturation marker CD83. DCs were capable of eliciting an HIV-1-antigen-specific response, as measured by expansion of autologous CD4(+) and CD8(+) T-cells. The expanded T-cells secreted gamma-IFN and interleukin (IL)-13, but not IL-10. The safety and immunogenicity of this DC vaccine are being evaluated in a Phase I/II clinical trial in chronically HIV-1-infected patients on HAART (clinicaltrials.gov identifier: NCT00796770).

Targeting concatenated HIV antigens to human CD40 expands a broad repertoire of multifunctional CD4+ and CD8+ T cells.

Objective:Targeting HIV antigens directly to dendritic cells using monoclonal antibodies against cell-surface receptors has been shown to evoke potent cellular immunity in animal models. The objective of this study was to configure an anti-human CD40 antibody fused to a string of five highly conserved CD4+ and CD8+ T-cell epitope-rich regions of HIV-1 Gag, Nef and Pol (&agr;CD40.HIV5pep), and then to demonstrate the capacity of this candidate therapeutic vaccine to target these HIV peptide antigens to human dendritic cells to expand functional HIV-specific T cells. Methods:Antigen-specific cytokine production using intracellular flow cytometry and multiplex bead-based assay, and suppression of viral inhibition, were used to characterize the T cells expanded by &agr;CD40.HIV5pep from HIV-infected patient peripheral blood mononuclear cell (PBMC) and dendritic cell/T-cell co-cultures. Results:This candidate vaccine expands memory CD4+ and CD8+ T cells specific to multiple epitopes within all five peptide regions across a wide range of major histocompatibility complex (MHC) haplotypes from HIV-infected patient PBMC and dendritic cell/T-cell co-cultures. These in vitro expanded HIV antigen-specific CD4+ and CD8+ T cells produce multiple cytokines and chemokines. &agr;CD40.HIV5pep-expanded CD8+ T cells have characteristics of cytotoxic effector cells and are able to kill autologous target cells and suppress HIV-1 replication in vitro. Conclusion:Our data demonstrate the therapeutic potential of this CD40-targeting HIV candidate vaccine in inducing a broad repertoire of multifunctional T cells in patients.

Delivering HIV Gagp24 to DCIR Induces Strong Antibody Responses In Vivo

Targeting dendritic cell-specific endocytic receptors using monoclonal antibodies fused to desired antigens is an approach widely used in vaccine development to enhance the poor immunogenicity of protein-based vaccines and to induce immune responses. Here, we engineered an anti-human DCIR recombinant antibody, which cross-reacts with the homologous cynomolgous macaque receptor and was fused via the heavy chain C-terminus to HIV Gagp24 protein (αDCIR.Gagp24). In vitro, αDCIR.Gagp24 expanded multifunctional antigen-specific memory CD4+ T cells recognizing multiple Gagp24 peptides from HIV-infected patient peripheral blood mononuclear cells. In non human primates, priming with αDCIR.Gagp24 without adjuvant elicited a strong anti-Gagp24 antibody response after the second immunization, while in the non-targeted HIV Gagp24 protein control groups the titers were weak. The presence of the double-stranded RNA poly(I:C) adjuvant significantly enhanced the anti-Gagp24 antibody response in all the groups and reduced the discrimination between the different vaccine groups. The avidity of the anti-Gagp24 antibody responses was similar with either αDCIR.Gagp24 or Gagp24 immunization, but increased from medium to high avidity in both groups when poly(I:C) was co-administered. This data provides a comparative analysis of DC-targeted and non-targeted proteins for their capacity to induce antigen-specific antibody responses in vivo. This study supports the further development of DCIR-based DC-targeting vaccines for protective durable antibody induction, especially in the absence of adjuvant.

P19-45. Development of a therapeutic HIV vaccine comprised of autologous dendritic cells loaded with a mixture of lipopeptide HIV antigens

Background Clinical trials have shown that dendritic cell (DC)-based vaccines can induce antigen-specific immune responses as well as clinical responses in patients with stage IV cancer. In this study, a monocyte-derived DC vaccine pulsed with HIV antigen lipopeptides (LIPO5) was developed in preparation for a pilot vaccine clinical trial (DALIA) aimed at boosting the cellular immune response in chronic HIV infected patients on highly-active antiretroviral therapy (HAART).

Vaccination with dendritic cells loaded with HIV-1 lipopeptides elicits broad T cell immunity and control of viral load in HIV infected patients

Background The DALIA trial tested the hypothesis that immunization with HIV peptide loaded Dendritic Cells (DC) may improve HIV immune responses and help to contain viral replication.

P16-49. Broad types of cytokines secreted by Gag-specific T cells from HIV infected patients on HAART

Background Previously described highly immunogenic epitopes from Gag are able to stimulate Th1/CTL cells. This response is essential for controlling progression of HIV infection. Techniques like IFN-γ-ELISPOT and IFN-γ intracellular staining are currently the standard methods to define antigen-specific immune responses. A recent method, the Luminex analysis allows the identification of responses characteristic of T cell subtypes through the secretion of not only IFN-γ but other key cytokines (IL-5, IL-13, IL-10, IL-21 and IL-17).

P16-29. HIV Nef-specific T cells: Th1/CTL, Th2 and Th17 responses

Background Identification of promiscuous antigenic regions is crucial for the design of an epitope-based vaccine. Still, the quality of the responses has been under estimated in most cases when only IFN-γ is used to measure anti-HIV cellular immunity. Luminex multiplexing system allows the identification of T cell responses characteristic of T cell subtypes through their secretion of not only Th1/CTL but other important sets of cytokines, including IL-5, Il-10, Il13, IL-21 and IL-17.