HIV-Tat immunization induces cross-clade neutralizing antibodies and CD4(+) T cell increases in antiretroviral-treated South African volunteers: a randomized phase II clinical trial.

Background: Although combined antiretroviral therapy (cART) has saved millions of lives, it is incapable of full immune reconstitution and virus eradication. The transactivator of transcription (Tat) protein is a key human immunodeficiency virus (HIV) virulence factor required for virus replication and transmission. Tat is expressed and released extracellularly by infected cells also under cART and in this form induces immune dysregulation, and promotes virus reactivation, entry and spreading.

Development of a novel AIDS vaccine: the HIV-1 transactivator of transcription protein vaccine.

Introduction: Classical approaches aimed at targeting the HIV-1 envelope as well as other structural viral proteins have largely failed. The HIV-1 transactivator of transcription (Tat) is a key HIV virulence factor, which plays pivotal roles in virus gene expression, replication, transmission and disease progression. Notably, anti-Tat Abs are uncommon in natural infection and, when present, correlate with the asymptomatic state and lead to lower or no disease progression.

HIV-1 Tat immunization restores immune homeostasis and attacks the HAART-resistant blood HIV DNA: results of a randomized phase II exploratory clinical trial.

Background: The phase II multicenter, randomized, open label, therapeutic trial (ISS T-002, Clinicaltrials.gov NCT00751595) was aimed at evaluating the immunogenicity and the safety of the biologically active HIV-1 Tat protein administered at 7.5 or 30 μg, given 3 or 5 times monthly, and at exploring immunological and virological disease biomarkers.

Surface-bound Tat inhibits antigen-specific CD8+ T-cell activation in an integrin-dependent manner.

Objective: The identification of still unrevealed mechanisms affecting the anti-HIV CD8 T-cell response in HIV-1 infection.

Design: Starting from the observation that anti-Tat immunization is associated with improved CD8 T-cell immunity, we developed both in-vitro and ex-vivo assays to characterize the effects of extra-cellular Tat on the adaptive CD8 T-cell response.

Methods: The effects of Tat on CD8 T-cell activation were assayed using CD8 T-cell clones specific for either cellular (MART-1) or viral (HIV-1 Nef) antigens, and HIV-1 Gag-specific CD8 T cells from HIV-1 patients.

The presence of anti-Tat antibodies in HIV-infected individuals is associated with containment of CD4+ T-cell decay and viral load, and with delay of disease progression: results of a 3-year cohort study.

Background: Tat is a key HIV-1 virulence factor, which plays pivotal roles in virus gene expression, replication, transmission and disease progression. After release, extracellular Tat accumulates in tissues and exerts effects on both the virus and the immune system, promoting immune activation and virus spreading while disabling the host immune defense. In particular, Tat binds Env spikes on virus particles forming a virus entry complex, which favors infection of dendritic cells and efficient transmission to T cells via RGD-binding integrins.

HIV-1 tat promotes integrin-mediated HIV transmission to dendritic cells by binding Env spikes and competes neutralization by anti-HIV antibodies.

Use of Env in HIV vaccine development has been disappointing. Here we show that, in the presence of a biologically active Tat subunit vaccine, a trimeric Env protein prevents in monkeys virus spread from the portal of entry to regional lymph nodes. This appears to be due to specific interactions between Tat and Env spikes that form a novel virus entry complex favoring R5 or X4 virus entry and productive infection of dendritic cells (DCs) via an integrin-mediated pathway.

Communication, recruitment and enrolment in the preventative and therapeutic phase I clinical trial against HIV/AIDS based on the recombinant HIV-1 Tat protein.

The role of volunteer recruitment in HIV vaccine trials has recently been considered particularly with respect to critical issues, such as motivation, psychological assessment and social impact. The preventative and therapeutic phase I trials based on the recombinant biologically active Tat vaccine candidate, sponsored in Italy by the Istituto Superiore di Sanità, included a specific centralised procedure (SCP) developed to support both the sponsor and the volunteers during trial enrolment and conduction. This process, which is an integrated, multidisciplinary, biomedical and psycho-socio-behavioural network, represented a novel and important aspect for the conduction and success of the clinical study.

Therapeutic immunization with HIV-1 Tat reduces immune activation and loss of regulatory T-cells and improves immune function in subjects on HAART.

Unlabelled: 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.

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.

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.

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.

Analysis of nickel-binding peptides in a human hepidermoid cancer cell line by Ni-NTA affinity chromatography and mass spectrometry.

To elucidate whether eukaryotic elongation factor 1A (eEF-1A) in a human hepidermoid cancer cell line (H1355) belonged to the family of the Ni-interacting protein, we analyzed the sequence of peptides obtained by on-Ni-NTA-agarose tryptic digestion of proteins from H1355 cell extract. LC/MS analysis showed the presence of several peptides mainly from abundant cellular proteins corresponding to eEF-1A, tubulin and actin. The results indicated that F-actin strongly binds to Ni-NTA-agarose whereas the other proteins are indirectly bound to the resin because of the formation of a protein-protein complex with actin.

Probing the secondary structure of a recombinant neuronal adaptor protein and its phosphotyrosine binding domains.

Rat brain Fe65 and its truncated forms corresponding to the combined PTB1 and PTB2 domains, as well as to the isolated PTB2 domain, were expressed in Escherichia coli and purified from inclusion bodies by affinity chromatography. The recombinant proteins were refolded and judged functionally active by their ability to interact with native APP. Limited proteolysis of recombinant Fe65 and PTB1-2 with trypsin, chymotrypsin and V8 proteases showed that the most sensitive proteoltytic sites were positioned at the level of the interdomain regions comprised between WW/PTB1 and PTB1/PTB2.

Elongation factor Ts from the Antarctic eubacterium Pseudoalteromonas haloplanktis TAC 125: biochemical characterization and cloning of the encoding gene.

The elongation factor Ts was isolated from the psychrophilic Antarctic eubacterium Pseudoalteromonas haloplanktis TAC 125 strain (PhEF-Ts), and its functional properties were studied. At 0 degrees C PhEF-Ts enhanced the [(3)H]GDP/GDP exchange rate on the preformed PhEF-Tu.[(3)H]GDP complex by 2 orders of magnitude even at very low Tu:Ts ratio, by lowering the energy of activation of the exchange reaction.

A long acidic domain affects the chromatographic behaviour of a neuronal adaptor protein on DEAE-Sepharose.

The stepwise chromatographic behaviour on DEAE-Sepharose of rat Fe65, a neuronal protein, was tested, using as eluants KCl, CaCl2, and MgCl2. Assays by western blot showed that Fe65 was eluted by CaCl2, at a ionic strength 20% lower than that of MgCl2 or KCl. Interestingly, in the case of a truncated Fe65, lacking a glutamic acid rich region at the N-terminus, the ionic strengths of the various eluants were almost identical.

Cloning, expression and evolution of the gene encoding the elongation factor 1alpha from a low thermophilic Sulfolobus solfataricus strain.

The gene encoding the elongation factor 1alpha (EF-1alpha) from the archaeon Sulfolobus solfataricus strain MT3 (optimum growth temperature 75 degrees C) was cloned, sequenced and expressed in Escherichia coli. The structural and biochemical properties of the purified enzyme were compared to those of EF-1alpha isolated from S. solfataricus strain MT4 (optimum growth temperature 87 degrees C).