Delipidated retroviruses as potential autologous therapeutic vaccines--a pilot experiment.

This pilot experiment in a simian immunodeficiency virus (SIV) chronic infection model aimed at extending our previous findings that vaccination with delipidated SIV resulted in more potent and diversified antiviral responses (1). Macaques chronically infected with SIVmac239 treated with antiretroviral therapy (ART) were vaccinated with autologous delipidated virus via consecutive lymph node targeted immunizations-1, 1 and 10 mug of virus spaced monthly. Results showed all animals had lasting viral load reduction approaching 1 log compared to set-point, and disease delay. Delipidation may enhance processing/ presentation of viral antigen eliciting potent antiviral control even at such late infection stage.

Simian/human immunodeficiency virus(89.6) expressing the chemokine genes MIP-1alpha, RANTES, or lymphotactin.

We constructed replication competent, attenuated, nef-deleted SHIV(89.6) that express the rhesus macaque chemokine genes MIP-1alpha, RANTES, or LTN from the nef region. The chemokine inserts were stable during several passages in CEMx174 cells and the viruses grew well in activated rhesus PBMC. Expression of virally encoded MIP-1alpha, RANTES, or LTN was detected in culture fluids from infected HOS CD4(+) CXCR4(+) cells, that were used because they have a low background production of these chemokines. The in vitro growth kinetics of all nef-deleted SHIV(89.6) were slower than the parental strain in both CEMx174 cells and rhesus PBMC. Rhesus macaques were susceptible to SHIV(89.6-MIP-1alpha), SHIV(89.6-RANTES), SHIV(89.6-LTN), and nef-deleted control SHIV(89.6-dLTN) infection via the intrarectal route using standard virus doses, and intact viruses were reisolated from infected animals throughout the interval of acute infection. SHIV expressing the chemokine genes MIP-1alpha, RANTES, or LTN may help determine the in vivo roles for these chemokines in modulating virus replication and disease.

Effects of virus burden and chemokine expression on immunity to SHIV in nonhuman primates.

HIV-1 vaccine candidates are designed to elicit Type 1 immune responses, including cytotoxic T cells and neutralizing antibodies. The type of immune response is influenced by many factors, including the levels of antigen expression and production of cytokines or chemokines; we designed a nonhuman primate study to evaluate the influence of these factors on protective immunity. Recombinant SHIV were engineered to express macrophage inflammatory protein-1 alpha (MIP-1alpha), regulated upon activation, normal T-cell expressed and secreted (RANTES), or Lymphotactin (Ltn) in place of nef in SHIV(89.6) (SHIV(89.6-MIP-1), SHIV(89.6-RANTES), SHIV(89.6-Ltn)). The parental virus SHIV(89.6) was included because it replicates to higher titer while still not causing disease. Control groups included animals that received a recombinant SHIV with a truncated chemokine construct (SHIV(89.6-dLtn)) and unvaccinated macaques. After pathogenic challenge with SHIV(89.6pd), animals from groups that received recombinant (nef-deleted) viruses had peak viremia levels three orders of magnitude lower than unvaccinated controls and increased survival times. Animals that received the original SHIV(89.6) (nef+) were highly resistant to both intrarectal and intravenous challenge with SHIV(89.6PD), and showed no signs of disease. There were no differences in survival times comparing unvaccinated and SHIV(89.6-dLtn) (control) groups, indicating that nef deleted viruses did not provide durable protection in this model. Strongest protection was seen in animals with the highest replicating virus (SHIV(89.6)), and the lower effect on survival after SHIV(89.6) nef-deleted vaccination, likely reflects differences in replication capacity. The protective effect of nef-deleted virus was partly restored by expressing Type 1 chemokines to augment viral immunity.

Primary African HIV clade A and D isolates: effective cross-clade neutralization with a quadruple combination of human monoclonal antibodies raised against clade B.

We investigated the ability of several human neutralizing monoclonal antibodies (nmAbs), originally raised against human immunodeficiency virus (HIV) clade B isolates, to neutralize primary clade A and D isolates as single agents and in combinations. All four primary HIV clade A isolates and five primary HIV clade D isolates tested were neutralized >99% by the quadruple combination of nmAbs IgG1b12, 2G12, 2F5, and 4E10. These mAbs recognize conserved epitopes on HIV-1 envelope (Env), resulting in strong cross-clade neutralization. Previously, we showed synergistic neutralization of primary HIV-1 clade C isolates in vitro by the same nMAb combination. We and others also showed neutralization of primary HIV clade B strains. Together, our data show that the quadruple combination of mAbs effectively neutralized primary HIV clade A, B, C, and D isolates.

Enhancement of cell mediated immune responses using lipid depleted lentivirus as immunogen: a novel approach for inducing recognition of new viral epitopes.

We tested the hypothesis that removal of viral lipids using diisopropylether can enhance antigenicity of SIVmac251. DIPE delipidation removed cholesterol from SIVmac251 without significant loss of viral protein or RNA. Mice immunized with the same SIV preparation but boosted with delipidated SIVmac251 exhibited significantly broader and higher cellular and humoral immune responses compared to live or AT-2-inactivated virus. As little as 1microg (total protein) of delipidated virus was sufficient to induce such enhanced immune responses. Thus, solvent treated lentivirus may provide a novel strategy to generate immune responses to additional viral epitopes.

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.