A DNA Vaccine Encoding Multiple HIV CD4 Epitopes Elicits Vigorous Polyfunctional, Long-Lived CD4+ and CD8+ T Cell Responses

Long-Lived CD4+ and CD8+ T Cell Responses. PLoS ONE 6(2): e16921. doi:10.1371/journal.pone.0016921 A DNA Vaccine Encoding Multiple HIV CD4 Epitopes + Elicits Vigorous Polyfunctional, Long-Lived CD4 and + CD8 T Cell Responses Daniela Santoro Rosa 0 Susan Pereira Ribeiro 0 Rafael Ribeiro Almeida 0 Eliane Conti Mairena 0 Edilberto Posto l 0 Jorge Kalil 0 Edecio Cunha-Neto 0 Lishomwa Ndhlovu, University of California San Francisco, United States of America 0 1 Laboratory of Clinical Immunology and Allergy-LIM60, Division of Clinical Immunology and Allergy, Department of Medicine, University of Sa o Paulo School of Medicine, Sa o Paulo, Brazil, 2 Heart Institute (InCor), University of Sa o Paulo School of Medicine, Sa o Paulo, Brazil, 3 Institute for Investigation in Immunology-INCT, Sa o Paulo, Brazil, 4 Division of Immunology-Federal University of Sa o Paulo-UNIFESP , Sa o Paulo , Brazil T-cell based vaccines against HIV have the goal of limiting both transmission and disease progression by inducing broad and functionally relevant T cell responses. Moreover, polyfunctional and long-lived specific memory T cells have been associated to vaccine-induced protection. CD4+ T cells are important for the generation and maintenance of functional CD8+ cytotoxic T cells. We have recently developed a DNA vaccine encoding 18 conserved multiple HLA-DR-binding HIV-1 CD4 epitopes (HIVBr18), capable of eliciting broad CD4+ T cell responses in multiple HLA class II transgenic mice. Here, we evaluated the breadth and functional profile of HIVBr18-induced immune responses in BALB/c mice. Immunized mice displayed high-magnitude, broad CD4+/CD8+ T cell responses, and 8/18 vaccine-encoded peptides were recognized. In addition, HIVBr18 immunization was able to induce polyfunctional CD4+ and CD8+ T cells that proliferate and produce any two cytokines (IFNc/TNFa, IFNc/IL-2 or TNFa/IL-2) simultaneously in response to HIV-1 peptides. For CD4+ T cells exclusively, we also detected cells that proliferate and produce all three tested cytokines simultaneously (IFNc/TNFa/IL-2). The vaccine also generated long-lived central and effector memory CD4+ T cells, a desirable feature for T-cell based vaccines. By virtue of inducing broad, polyfunctional and long-lived T cell responses against conserved CD4+ T cell epitopes, combined administration of this vaccine concept may provide sustained help for CD8+ T cells and antibody responses- elicited by other HIV immunogens. - Funding: This research was supported by the Brazilian National Research Council (CNPq), Sao Paulo State Research Funding Agency (FAPESP), International Centre of Genetic Engineering and Biotechnology (ICGEB) and by the Brazilian Ministry of Health (Brazil). D. S. Rosa, S. P. Ribeiro and R. R. Almeida are recipients of a Sao Paulo State Research Funding Agency (FAPESP) fellowship. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. The International Centre of Genetic Engineering and Biotechnology (ICGEB) is a non-profit organization related to the UNESCO which funds basic research in developing and middle-income countries. There are no links (employment, consultancy, patents, products in development or marketed products) between the authors and the ICGEB that may alter the authors adherence to all the PLoS ONE policies on sharing data and materials. The use of the peptide combination for vaccination purposes, among others, has been patented (international application number PCT/BR2006/000175). . These authors contributed equally to this work. Despite the success of antiretroviral treatment, a safe and effective HIV vaccine is the most promising strategy for controlling the AIDS pandemic, especially in developing countries. HIV vaccine strategies that focus on the generation of virus-specific Tcell responses have the goal of limiting both transmission and disease progression by controlling HIV viral loads [1]. To date, two efficacy trials assessed HIV-specific cellular mediated immunity. The STEP vaccine trial developed by Merck used a replication-defective Ad5 vector, expressing Gag, Pol and Nef proteins from HIV-1 [2]. The results from this trial demonstrated that it neither prevented HIV-1 infection nor reduced viral load in subsequently infected subjects [3,4]. Immunological analyses revealed that each vaccinated individual recognized an average of only three epitopes [5]. The narrowness of the induced immune responses may have been an important factor in the lack of vaccine efficacy [3]. Indeed, non-human primate studies have shown that vaccines that induced broad CD8+ and CD4+ T cell responses can control peak SIV viremia [6]. In the recently reported RV144 HIV-1 vaccine trial conducted in Thailand, an immunization strategy based on recombinant canarypox priming followed by a protein boosting generated modest protection against the acquisition of HIV infection. Immunological analysis of samples from the study showed that the vaccine-induced immune response was essentially composed of CD4+ T cells and binding antibodies; no IFNc or IL-2 secreting HIV-specific CD8+ T cells were detected [7]. However, the same immunogens induced cytotoxic immune responses in a minority of vaccinees in previous studies [8]. At any event, data from the RV 144 trial supported the notion that CD4+ T cells could play a protective role in HIV vaccine-induced immunity. CD4+ T cells can contribute to protection against viral infection by both indirect and direct manners [911]. CD4+ T cell can help induce and maintain CD8+ and B cell responses. The main contribution of CD4+ T cells is to provide help to full differentiation and maintenance of cytotoxic CD8+ T cells and B cells. They promote the generation of CD8+ cytotoxic T cell response (CTL) able to control viral replication [1214] as well as mobilization of CTLs to peripheral sites of infection [15]. Furthermore, CD4+ T cells can promote B cell differentiation into plasma cells to produce neutralizing antibodies and assist memory B cells responses to re-infection [16]. CD4+ T cells can also exert direct and indirect antiviral effects in retroviral infection. The outcome of retroviral infection depends on the magnitude and duration of virus-specific CD4+T cell responses [17]. A direct antiviral effect of CD4+ T cells was also observed in SIV infection. CD4+ T cells induce apoptosis of SIV-infected macrophages [18]. The presence of SIV-specific CD4+ T cell responses with a cytotoxic phenotype was associated with the control of rebounding viremia in CD8+ depleted SIV-infected macaques [19]. Further in support of a protective role for CD4+ T cell responses, it has been shown that elite controller SIV-infected macaques mount broad CD4+-specific T cell responses, and that certain macaque class II alleles are associated with significantly decreased viral (...truncated)