Antonella Tripiciano

The Presence of Anti-Tat Antibodies Is Predictive of Long-Term Nonprogression to AIDS or Severe Immunodeficiency: Findings in a Cohort of HIV-1 Seroconverters

The human immunodeficiency virus (HIV) type 1 Tat protein plays a key role in the life cycle of the virus and in pathogenesis and is highly conserved among HIV subtypes. On the basis of this and of safety, immunogenicity, and efficacy findings in monkeys, Tat is being tested as a vaccine in phase 1 trials. Here, we evaluated the incidence and risk of progression to advanced HIV disease by anti-Tat serostatus in a cohort of 252 HIV-1 seroconverters. The risk of progression was lower in the anti-Tat-positive subjects than in the anti-Tat-negative subjects. Progression was faster in the persistently anti-Tat-negative subjects than in the transiently anti-Tat-positive subjects, and no progression was observed in the persistently anti-Tat-positive subjects.

Therapeutic immunization with hiv-1 tat reduces immune activation and loss of regulatory t-cells and improves immune function in subjects on HAART

Although HAART suppresses HIV replication, it is often unable to restore immune homeostasis. Consequently, non-AIDS-defining diseases are increasingly seen in treated individuals. This is attributed to persistent virus expression in reservoirs and to cell activation. Of note, in CD4+ T cells and monocyte-macrophages of virologically-suppressed individuals, there is continued expression of multi-spliced transcripts encoding HIV regulatory proteins. Among them, Tat is essential for virus gene expression and replication, either in primary infection or for virus reactivation during HAART, when Tat is expressed, released extracellularly and exerts, on both the virus and the immune system, effects that contribute to disease maintenance. Here we report results of an ad hoc exploratory interim analysis (up to 48 weeks) on 87 virologically-suppressed HAART-treated individuals enrolled in a phase II randomized open-label multicentric clinical trial of therapeutic immunization with Tat (ISS T-002). Eighty-eight virologically-suppressed HAART-treated individuals, enrolled in a parallel prospective observational study at the same sites (ISS OBS T-002), served for intergroup comparison. Immunization with Tat was safe, induced durable immune responses, and modified the pattern of CD4+ and CD8+ cellular activation (CD38 and HLA-DR) together with reduction of biochemical activation markers and persistent increases of regulatory T cells. This was accompanied by a progressive increment of CD4+ T cells and B cells with reduction of CD8+ T cells and NK cells, which were independent from the type of antiretroviral regimen. Increase in central and effector memory and reduction in terminally-differentiated effector memory CD4+ and CD8+ T cells were accompanied by increases of CD4+ and CD8+ T cell responses against Env and recall antigens. Of note, more immune-compromised individuals experienced greater therapeutic effects. In contrast, these changes were opposite, absent or partial in the OBS population. These findings support the use of Tat immunization to intensify HAART efficacy and to restore immune homeostasis. Trial registration ClinicalTrials.gov NCT00751595

Sequence Conservation and Antibody Cross-Recognition of Clade B Human Immunodeficiency Virus (HIV) Type 1 Tat Protein in HIV-1–Infected Italians, Ugandans, and South Africans

We determined immune cross-recognition and the degree of Tat conservation in patients infected by local human immunodeficiency virus (HIV) type 1 strains. The data indicated a similar prevalence of total and epitope-specific anti-Tat IgG in 578 serum samples from HIV-infected Italian (n=302), Ugandan (n=139), and South African (n=137) subjects, using the same B clade Tat protein that is being used in vaccine trials. In particular, anti-Tat antibodies were detected in 13.2%, 10.8%, and 13.9% of HIV-1-infected individuals from Italy, Uganda, and South Africa, respectively. Sequence analysis results indicated a high similarity of Tat from the different circulating viruses with BH-10 Tat, particularly in the 1-58 amino acid region, which contains most of the immunogenic epitopes. These data indicate an effective cross-recognition of a B-clade laboratory strain-derived Tat protein vaccine by individuals infected with different local viruses, owing to the high similarity of Tat epitopes.

Efficient mucosal delivery of the HIV‐1 Tat protein using the synthetic lipopeptide MALP‐2 as adjuvant

A major requirement for HIV/AIDS research is the development of a mucosal vaccine that stimulates humoral and cell‐mediated immune responses at systemic and mucosal levels, thereby blocking virus replication at the entry port. Thus, a vaccine prototype based on biologically active HIV‐1 Tat protein as antigen and the synthetic lipopeptide, macrophage‐activating lipopeptide‐2 (MALP‐2), asa mucosal adjuvant was developed. Intranasal administration to mice stimulated systemic and mucosal anti‐Tat antibody responses, and Tat‐specific T cell responses, that were more efficient than those observed after i.p. immunization with Tat plus incomplete Freunds adjuvant. Major linear B cell epitopes mapped within aa 1–20 and 46–60, whereas T cell epitopes were identified within aa 36–50 and 56–70. These epitopes have also been described in vaccinated primates and in HIV‐1‐infected individuals with better prognosis. Analysis of the anti‐Tat IgG isotypes in serum, and the cytokine profile of spleen cells indicated that a dominant Th1 helper response was stimulated by Tat plus MALP‐2, as opposed to the Th2 response observed with Tat plus incomplete Freunds adjuvant. Tat‐specific IFN‐γ‐producing cells were significantly increased only in response to Tat plus MALP‐2. These data suggest that Malp‐2 may represent an optimal mucosal adjuvant for candidate HIV vaccines based on Tat alone or in combination with other HIV antigens.

HIV-1 Tat addresses dendritic cells to induce a predominant Th1-type adaptive immune response that appears prevalent in the asymptomatic stage of infection

Tat is an early regulatory protein that plays a major role in human HIV-1 replication and AIDS pathogenesis, and therefore, it represents a key target for the host immune response. In natural infection, however, Abs against Tat are produced only by a small fraction (∼20%) of asymptomatic individuals and are rarely seen in progressors, suggesting that Tat may possess properties diverting the adaptive immunity from generating humoral responses. Here we show that a Th1-type T cell response against Tat is predominant over a Th2-type B cell response in natural HIV-1 infection. This is likely due to the capability of Tat to selectively target and very efficiently enter CD1a-expressing monocyte-derived dendritic cells (MDDC), which represent a primary target for the recognition and response to virus Ag. Upon cellular uptake, Tat induces MDDC maturation and Th1-associated cytokines and β-chemokines production and polarizes the immune response in vitro to the Th1 pattern through the transcriptional activation of TNF-α gene expression. This requires the full conservation of Tat transactivation activity since neither MDDC maturation nor TNF-α production are found with either an oxidized Tat, which does not enter MDDC, or with a Tat protein mutated in the cysteine-rich region (cys22 Tat), which enters MDDC as the wild-type Tat but is transactivation silent. Consistently with these data, inoculation of monkeys with the native wild-type Tat induced a predominant Th1 response, whereas cys22 Tat generated mostly Th2 responses, therefore providing evidence that Tat induces a predominant Th1 polarized adaptive immune response in the host.

Phase I therapeutic trial of the HIV-1 Tat protein and long term follow-up

A randomized, double blind, placebo-controlled phase I vaccine trial based on the native Tat protein was conducted in HIV-infected asymptomatic individuals. The vaccine was administered five times subcute with alum or intradermally without adjuvant at 7.5microg, 15microg or 30microg doses, respectively. The Tat vaccine was well tolerated both locally and systemically and induced and/or maintained Tat-specific T helper (Th)-1 T-cell responses and Th-2 responses in all subjects with a wide spectrum of functional anti-Tat antibodies, rarely seen in HIV-infected subjects. The data indicate the achievement of both the primary (safety) and secondary (immunogenicity) endpoints of the study.

The preventive phase I trial with the HIV-1 Tat-based vaccine.

The native HIV-1 Tat protein was chosen as vaccine candidate for phase I clinical trials based on its role in the natural infection and AIDS pathogenesis, on the association of Tat-specific immune response with the asymptomatic stage as well as on its sequence conservation among HIV clades. A randomized, double blind, placebo-controlled phase I study (ISS P-001) was conducted in healthy adult volunteers without identifiable risk of HIV infection. Tat was administered 5 times monthly, subcute in alum or intradermic alone at 7.5 microg, 15 microg or 30 microg, respectively (ClinicalTrials.gov identifier: NCT00529698). Vaccination with Tat resulted to be safe and well tolerated (primary endpoint) both locally and systemically. In addition, Tat induced both Th1 and Th2 type specific immune responses in all subjects (secondary endpoint) with a wide spectrum of functional antibodies that are rarely seen in natural infection, providing key information for further clinical development of the Tat vaccine candidate.

HIV-1 Tat-Based Vaccines: From Basic Science to Clinical Trials

Vaccination against human immunodeficiency virus (HIV)-1 infection requires candidate antigen(s) (Ag) capable of inducing an effective, broad, and long-lasting immune response against HIV-1 despite mutation events leading to differences in virus clades. The HIV-1 Tat protein is more conserved than envelope proteins, is essential in the virus life cycle and is expressed very early upon virus entry. In addition, both humoral and cellular responses to Tat have been reported to correlate with a delayed progression to disease in both humans and monkeys. This suggested that Tat is an optimal target for vaccine development aimed at controlling virus replication and blocking disease onset. Here are reviewed the results of our studies including the effects of the Tat protein on monocyte-derived dendritic cells (MDDCs) that are key antigen-presenting cells (APCs), and the results from vaccination trials with both the Tat protein or tat DNA in monkeys. We provide evidence that the HIV-1 Tat protein is very efficiently taken up by MDDCs and promotes T helper (Th)-1 type immune responses against itself as well as other Ag. In addition, a Tat-based vaccine elicits an immune response capable of controlling primary infection of monkeys with the pathogenic SHIV89.6P at its early stages allowing the containment of virus spread. Based on these results and on data of Tat conservation and immune cross-recognition in field isolates from different clades, phase I clinical trials are being initiated in Italy for both preventive and therapeutic vaccination.

Persistent Immune Responses Induced by a Human Immunodeficiency Virus DNA Vaccine Delivered in Association with Electroporation in the Skin of Nonhuman Primates

Strategies to improve vaccine efficacy are still required, especially in the case of chronic infections, including human immunodeficiency virus (HIV). DNA vaccines have potential advantages over conventional vaccines; however, low immunological efficacy has been demonstrated in many experiments involving large animals and in clinical trials. To improve the immunogenicity of DNA vaccines, we have designed a plasmid vector exploiting the binding capacity of the bovine papillomavirus E2 protein and we have used electroporation (EP) to increase DNA uptake after intradermal inoculation. We demonstrated, in nonhuman primates (NHPs), efficient induction of anti-HIV immunity with an improved DNA vaccine vector encoding an artificial fusion protein, consisting of several proteins and selected epitopes from HIV-1. We show that a DNA vaccine delivery method combining intradermal injection and noninvasive EP dramatically increased expression of the vaccine antigen selectively in the epidermis, and our observations strongly suggest the involvement of Langerhans cells in the strength and quality of the anti-HIV immune response. Although the humoral responses to the vaccine were transient, the cellular responses were exceptionally robust and persisted, at high levels, more than 2 years after the last vaccine boost. The immune responses were characterized by the induction of significant proportions of T cells producing both interferon-gamma and interleukin-2 cytokines, in both subpopulations, CD4(+) and CD8(+). This strategy is an attractive approach for vaccination in humans because of its high efficacy and the possible use of newly developed devices for EP.

HIV-1 Tat-Based Vaccines: An Overview and Perspectives in the Field of HIV/AIDS Vaccine Development

The HIV epidemic continues to represent one of the major problems worldwide, particularly in the Asia and Sub-Saharan regions of the world, with social and economical devastating effects. Although antiretroviral drugs have had a dramatically beneficial impact on HIV-infected individuals that have access to treatment, it has had a negligible impact on the global epidemic. Hence, the inexorable spreading of the HIV pandemic and the increasing deaths from AIDS, especially in developing countries, underscore the urgency for an effective vaccine against HIV/AIDS. However, the generation of such a vaccine has turned out to be extremely challenging. Here we provide an overview on the rationale for the use of non-structural HIV proteins, such as the Tat protein, alone or in combination with other HIV early and late structural HIV antigens, as novel, promising preventative and therapeutic HIV/AIDS vaccine strategies.