Flavia Ferrantelli

Postnatal Passive Immunization of Neonatal Macaques with a Triple Combination of Human Monoclonal Antibodies against Oral Simian-Human Immunodeficiency Virus Challenge

ABSTRACT To develop prophylaxis against mother-to-child human immunodeficiency virus (HIV) transmission, we established a simian-human immunodeficiency virus (SHIV) infection model in neonatal macaques that mimics intrapartum mucosal virus exposure (T. W. Baba et al., AIDS Res. Hum. Retroviruses 10:351–357, 1994). Using this model, neonates were protected from mucosal SHIV-vpu+challenge by pre- and postnatal treatment with a combination of three human neutralizing monoclonal antibodies (MAbs), F105, 2G12, and 2F5 (Baba et al., Nat. Med. 6:200–206, 2000). In the present study, we used this MAb combination only postnatally, thereby significantly reducing the quantity of antibodies necessary and rendering their potential use in humans more practical. We protected two neonates with this regimen against oral SHIV-vpu+ challenge, while four untreated control animals became persistently infected. Thus, synergistic MAbs protect when used as immunoprophylaxis without the prenatal dose. We then determined in vitro the optimal MAb combination against the more pathogenic SHIV89.6P, a chimeric virus encodingenv of the primary HIV89.6. Remarkably, the most potent combination included IgG1b12, which alone does not neutralize SHIV89.6P. We administered the combination of MAbs IgG1b12, 2F5, and 2G12 postnatally to four neonates. One of the four infants remained uninfected after oral challenge with SHIV89.6P, and two infants had no or a delayed CD4+ T-cell decline. In contrast, all control animals had dramatic drops in their CD4+ T cells by 2 weeks postexposure. We conclude that our triple MAb combination partially protected against mucosal challenge with the highly pathogenic SHIV89.6P. Thus, combination immunoprophylaxis with passively administered synergistic human MAbs may play a role in the clinical prevention of mother-to-infant transmission of HIV type 1.

Complete Protection of Neonatal Rhesus Macaques against Oral Exposure to Pathogenic Simian-Human Immunodeficiency Virus by Human Anti-HIV Monoclonal Antibodies

Because milk-borne transmission of human immunodeficiency virus (HIV) diminishes the benefits of perinatal antiviral drug therapy in developing countries, we have developed a new strategy to prevent postnatal and, possibly, intrapartum virus transmission in a primate model. Eight neonatal rhesus macaques were exposed orally to pathogenic simian-human immunodeficiency virus (SHIV); 4 neonates were then given intramuscular postexposure prophylaxis with 3 anti-HIV human neutralizing monoclonal antibodies (nMAbs) with potent cross-clade and cross-group neutralization activity. Untreated infants experienced high viral RNA levels and CD4(+) T-cell losses and died (median survival time, 5.5 weeks). In contrast, all 4 nMAb-treated neonates were protected from infection (P=.028); their plasma, peripheral blood mononuclear cells, and lymph nodes remained virus negative for >1 year. These data are important for designing clinical trials in human neonates and have general implications for AIDS vaccine development, as the epitopes recognized by the 3 nMAbs are conserved among diverse primary isolates.

Neutralizing antibodies against HIV -- back in the major leagues?

The past few months have seen encouraging successes for neutralizing antibodies against HIV; human monoclonal antibodies targeting conserved HIV envelope epitopes potently neutralized primary virus isolates, including strains of different clades. In primates, passive immunization with combinations containing human monoclonal antibodies completely prevented infection, even after mucosal virus challenges. Epitopes recognized by the protective monoclonal antibodies are important determinants for protection and provide a rational basis for AIDS vaccine development.

Antibody protection: passive immunization of neonates against oral AIDS virus challenge

We have established models for intrapartum and milk-borne HIV transmission by orally challenging neonatal macaques with chimeric simian-human immunodeficiency viruses (SHIVs). This allowed us to test safety and efficacy of passive immunization with human neutralizing monoclonal antibodies (nmAbs), which had been isolated from HIV clade B-infected individuals and which target conserved, functionally important epitopes. The nmAbs studied were F105 or IgG1b12, b12 for short (directed against the CD4 binding site), 2G12 (anti-gp120), 2F5 and 4E10 (both anti-gp41). Out of 16 newborn macaques challenged orally with different SHIV strains, 11 were completely protected by triple or quadruple nmAb combinations, even by post-exposure prophylaxis. In vitro, the combination of b12, 2G12, 2F5 and 4E10 potently neutralized primary HIV isolates of clades A, B, C, and D. Our data suggest that passive immunization with currently available anti-HIV clade B nmAbs could play a role in preventing transmission of non-clade B isolates through breastfeeding. We furthermore conclude that the epitopes recognized by the nmAbs in our successful passive immunization studies are important determinants for protection and provide targets for developing neutralizing antibody-response-based, active AIDS vaccines.

Immunoprophylaxis to prevent mother-to-child transmission of HIV-1.

Antiretroviral therapy can profoundly reduce the risk of mother-to-child transmission (MTCT) of HIV, but the drugs have a relatively short half-life and should thus be administered throughout breast-feeding to optimally prevent postnatal infection of the infant. The potential toxicities and the development of resistance may limit the long-term efficacy of antiretroviral prophylaxis, and a safe and effective active/passive immunoprophylaxis regimen, begun at birth, and potentially overlapping with interpartum or neonatal chemoprophylaxis, would pose an attractive alternative. This review draws on data presented at the Ghent Workshop on prevention of breast milk transmission and on selected issues from a workshop specifically relating to immunoprophylaxis held in Seattle in October 2002. This purpose of this review is to address the scientific rationale for the development of passive (antibody) and active (vaccine) immunization strategies for prevention of MTCT. Data regarding currently or imminently available passive and active immunoprophylaxis products are reviewed for their potential use in neonatal trials within the coming 1-2 years.

Potent Cross-Group Neutralization of Primary Human Immunodeficiency Virus Isolates with Monoclonal Antibodies—Implications for Acquired Immunodeficiency Syndrome Vaccine

Human immunodeficiency virus type 1 (HIV-1) is phylogenetically classified into groups and clades (or subtypes). Human neutralizing monoclonal antibodies (nMAbs), originally isolated from individuals infected with HIV-1 group M-clade B, neutralized not only primary HIV-1 clade B isolates in vitro but also primary isolates of other group M clades (A, C, D, E, and F). This corrected the previously held notion that primary HIV-1 isolates are resistant to neutralizing antibodies. Here we show that anti-HIV-1 group M-clade B nMAbs potently neutralized primary isolates of the phylogenetically distant HIV-1 group O. We and others have previously shown that passive immunization with human nMAbs protected adult or neonatal primates against infection with simian-human immunodeficiency virus strains encoding HIV-1 group M-clade B envelope genes. The in vitro cross-group neutralization shown here underscores the broad potential of these nMAbs against divergent virus variants and the relevance of their epitopes in the design of acquired immunodeficiency syndrome vaccines.

T-tropic human immunodeficiency virus (HIV) type 1 Nef protein enters human monocyte-macrophages and induces resistance to HIV replication: a possible mechanism of HIV T-tropic emergence in AIDS.

Increasing interest has been devoted to the role that monocyte-macrophages play in the pathogenesis of AIDS. The hypothesis of an involvement in AIDS pathogenesis of human/simian immunodeficiency virus (HIV/SIV) Nef also is currently under evaluation by many investigators. The original basis of this hypothesis came from evidence that monkeys infected with a nef-deleted SIV strain failed to develop simian AIDS. Here, we show that treatment of human monocyte-derived macrophages (MDM) with recombinant HIV-1 Nef protein (rNef) induces a strong inhibition of the replication of either macrophage (M-) or dual-tropic HIV-1 strains. Through cytofluorimetric analyses, we detected internalization of FITC-conjugated rNef in MDM as early as 6 h after treatment. Confocal microscope observations demonstrated that the intracellular distribution of internalized rNef was identical to that of endogenously produced Nef. Down-regulation of the CD4 HIV receptor detected upon rNef treatment of MDM suggested that the rNef-induced HIV inhibition occurred at the virus entry step. This deduction was strengthened by the observation that CD4-independent infection was totally insensitive to rNef treatment. The specificity of all observed effects was demonstrated by immunodepletion of rNef. Finally, we showed that the resistance to HIV replication induced by rNef treatment in MDM favours the spread of T-tropic over M-tropic HIV strains in doubly infected CD4(+) lymphocyte-MDM co-cultures. We propose that extracellular Nef contributes to AIDS pathogenesis by inducing resistance to M-tropic HIV replication in MDM, thereby facilitating the switching from M- to T-tropic HIV prevalence that correlates frequently with AIDS progression.

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.

cis Expression of the F12 Human Immunodeficiency Virus (HIV) Nef Allele Transforms the Highly Productive NL4-3 HIV Type 1 to a Replication-Defective Strain: Involvement of both Env gp41 and CD4 Intracytoplasmic Tails

ABSTRACT F12 human immunodeficiency virus type 1 (HIV-1) nef is a naturally occurring nef mutant cloned from the provirus of a nonproductive, nondefective, and interfering HIV-1 variant (F12-HIV). We have already shown that cells stably transfected with a vector expressing the F12-HIV nef allele do not downregulate CD4 receptors and, more peculiarly, become resistant to the replication of wild type (wt) HIV. In order to investigate the mechanism of action of such an HIV inhibition, the F12-HIVnef gene was expressed in the context of the NL4-3 HIV-1 infectious molecular clone by replacing the wt nef gene (NL4-3/chi). Through this experimental approach we established the following. First, NL4-3/chi and nef-defective (Δnef) NL4-3 viral particles behave very similarly in terms of viral entry and HIV protein production during the first replicative cycle. Second, no viral particles were produced from cells infected with NL4-3/chi virions, whatever the multiplicity of infection used. The viral inhibition apparently occurs at level of viral assembling and/or release. Third, this block could not be relieved by in-trans expression of wt nef. Finally, NL4-3/chi reverts to a producer HIV strain when F12-HIV Nef is deprived of its myristoyl residue. Through a CD4 downregulation competition assay, we demonstrated that F12-HIV Nef protein potently inhibits the CD4 downregulation induced by wt Nef. Moreover, we observed a redistribution of CD4 receptors at the cell margin induced by F12-HIV Nef. These observations strongly suggest that F12-HIV Nef maintains the ability to interact with the intracytoplasmic tail of the CD4 receptor molecule. Remarkably, we distinguished the intracytoplasmic tails of Env gp41 and CD4 as, respectively, viral and cellular targets of the F12-HIV Nef-induced viral retention. For the first time, the inhibition of the viral life cycle by means of in-cis expression of a Nef mutant is here reported. Delineation of the F12-HIV Nef mechanism of action may offer additional approaches to interference with the propagation of HIV infection.

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. These Tat effects do not require Tat-transactivation activity and are blocked by anti-integrin antibodies (Abs). Productive DC infection promoted by Tat is associated with a highly efficient virus transmission to T cells. In the Tat/Env complex the cysteine-rich region of Tat engages the Env V3 loop, whereas the Tat RGD sequence remains free and directs the virus to integrins present on DCs. V2 loop deletion, which unshields the CCR5 binding region of Env, increases Tat/Env complex stability. Of note, binding of Tat to Env abolishes neutralization of Env entry or infection of DCs by anti-HIV sera lacking anti-Tat Abs, which are seldom present in natural infection. This is reversed, and neutralization further enhanced, by HIV sera containing anti-Tat Abs such as those from asymptomatic or Tat-vaccinated patients, or by sera from the Tat/Env vaccinated monkeys. Thus, both anti-Tat and anti-Env Abs are required for efficient HIV neutralization. These data suggest that the Tat/Env interaction increases HIV acquisition and spreading, as a mechanism evolved by the virus to escape anti-Env neutralizing Abs. This may explain the low effectiveness of Env-based vaccines, which are also unlikely to elicit Abs against new Env epitopes exposed by the Tat/Env interaction. As Tat also binds Envs from different clades, new vaccine strategies should exploit the Tat/Env interaction for both preventative and therapeutic interventions.