AAV-Delivered Antibody Mediates Significant Protective Effects against SIVmac239 Challenge in the Absence of Neutralizing Activity

Long-term delivery of potent broadly-neutralizing antibodies is a promising approach for the prevention of HIV-1 infection. We used AAV vector intramuscularly to deliver anti-SIV monoclonal antibodies (mAbs) in IgG1 form to rhesus monkeys. Persisting levels of delivered mAb as high as 270 mug/ml were achieved. However, host antibody responses to the delivered antibody were observed in 9 of the 12 monkeys and these appeared to limit the concentration of delivered antibody that could be achieved. This is reflected in the wide range of delivered mAb concentrations that were achieved: 1-270 mug/ml. Following repeated, marginal dose, intravenous challenge with the difficult-to-neutralize SIVmac239, the six monkeys in the AAV-5L7 IgG1 mAb group showed clear protective effects despite the absence of detectable neutralizing activity against the challenge virus. The protective effects included: lowering of viral load at peak height; lowering of viral load at set point; delay in the time to peak viral load from the time of the infectious virus exposure. All of these effects were statistically significant. In addition, the monkey with the highest level of delivered 5L7 mAb completely resisted six successive SIVmac239 i.v. challenges, including a final challenge with a dose of 10 i.v. infectious units. Our results demonstrate the continued promise of this approach for the prevention of HIV-1 infection in people. However, the problem of anti-antibody responses will need to be understood and overcome for the promise of this approach to be effectively realized

Electron Tomography of the Contact between T Cells and SIV/HIV-1: Implications for Viral Entry

The envelope glycoproteins of primate lentiviruses, including human and simian immunodeficiency viruses (HIV and SIV), are heterodimers of a transmembrane glycoprotein (usually gp41), and a surface glycoprotein (gp120), which binds CD4 on target cells to initiate viral entry. We have used electron tomography to determine the three-dimensional architectures of purified SIV virions in isolation and in contact with CD4+ target cells. The trimeric viral envelope glycoprotein surface spikes are heterogeneous in appearance and typically ∼120 Å long and ∼120 Å wide at the distal end. Docking of SIV or HIV-1 on the T cell surface occurs via a neck-shaped contact region that is ∼400 Å wide and consistently consists of a closely spaced cluster of five to seven rod-shaped features, each ∼100 Å long and ∼100 Å wide. This distinctive structure is not observed when viruses are incubated with T lymphocytes in the presence of anti-CD4 antibodies, the CCR5 antagonist TAK779, or the peptide entry inhibitor SIVmac251 C34. For virions bound to cells, few trimers were observed away from this cluster at the virion–cell interface, even in cases where virus preparations showing as many as 70 envelope glycoprotein trimers per virus particle were used. This contact zone, which we term the “entry claw”, provides a spatial context to understand the molecular mechanisms of viral entry. Determination of the molecular composition and structure of the entry claw may facilitate the identification of improved drugs for the inhibition of HIV-1 entry

Highly Efficient Autologous HIV-1 Isolation by Coculturing Macrophage With Enriched CD4+ T Cells From HIV-1 Patients

We described a novel HIV autologous isolation method based in coculturing macrophages and CD4+T-cell-enriched fractions from peripheral blood collected from antiretroviral-treated (ART) HIV patients. This method allows the isolation of high viral titers of autologous viruses, over 1010HIV RNA copies/ml, and reduces the time required to produce necessary amounts for virus for use as antigens presented by monocyte-derived myeloid cells in HIV therapeutic vaccine approaches. By applying these high titer and autologous virus produced in the patient-derived cells, we intended to elicit a boost of the immunological system response in HIV therapeutic vaccines in clinical trials.This study was partially supported by grants from the Spanish Ministry of Economy (MINECO) (grants: SAF2015-66193-R, SAF-2017-88089-R, RTI2018-096309-B-I00); the Fondo Europeo para el Desarrollo Regional (FEDER); the SPANISH AIDS Research Network RD16/0025/0002 and RD16/0025/0014-ISCIII-FEDER (RIS); the Fondo de Investigación Sanitaria (FIS) PI12/01247 and PI20/00676; and HIVACAT program and the CERCA Programme/Generalitat de Catalunya SGR 615 and SGR 653. The project leading to these results has received funding from “la Caixa” Foundation under agreement. This study was also supported in part with federal funds from the National Cancer Institute, National Institutes of Health, under contract Nos. HHSN261200800001E and 75N91019D00024.S

Topically Applied Recombinant Chemokine Analogues Fully Protect Macaques from Vaginal Simian-Human Immunodeficiency Virus Challenge

Effective strategies for preventing human immunodeficiency virus infection are urgently needed, but recent failures in key clinical trials of vaccines and microbicides highlight the need for new approaches validated in relevant animal models. Here, we show that 2 new chemokine (C-C motif) receptor 5 inhibitors, 5P12-RANTES (regulated on activation, normal T cell expressed and secreted) and 6P4-RANTES, fully protect against infection in the rhesus vaginal challenge model. These highly potent molecules, which are amenable to low-cost production, represent promising new additions to the microbicides pipelin

ADCC Develops Over Time during Persistent Infection with Live-Attenuated SIV and Is Associated with Complete Protection against SIVmac251SIV_{mac}251 Challenge

Live-attenuated strains of simian immunodeficiency virus (SIV) routinely confer apparent sterilizing immunity against pathogenic SIV challenge in rhesus macaques. Understanding the mechanisms of protection by live-attenuated SIV may provide important insights into the immune responses needed for protection against HIV-1. Here we investigated the development of antibodies that are functional against neutralization-resistant SIV challenge strains, and tested the hypothesis that these antibodies are associated with protection. In the absence of detectable neutralizing antibodies, Env-specific antibody-dependent cell-mediated cytotoxicity (ADCC) emerged by three weeks after inoculation with SIVΔnef, increased progressively over time, and was proportional to SIVΔnef replication. Persistent infection with SIVΔnef elicited significantly higher ADCC titers than immunization with a non-persistent SIV strain that is limited to a single cycle of infection. ADCC titers were higher against viruses matched to the vaccine strain in Env, but were measurable against viruses expressing heterologous Env proteins. In two separate experiments, which took advantage of either the strain-specificity or the time-dependent maturation of immunity to overcome complete protection against $SIV_{mac}251$ challenge, measures of ADCC activity were higher among the SIVΔnef-inoculated macaques that remained uninfected than among those that became infected. These observations show that features of the antibody response elicited by SIVΔnef are consistent with hallmarks of protection by live-attenuated SIV, and reveal an association between Env-specific antibodies that direct ADCC and apparent sterilizing protection by SIVΔnef

ADCC Develops Over Time during Persistent Infection with Live-Attenuated SIV and Is Associated with Complete Protection against SIV(mac)251 Challenge

Live-attenuated strains of simian immunodeficiency virus (SIV) routinely confer apparent sterilizing immunity against pathogenic SIV challenge in rhesus macaques. Understanding the mechanisms of protection by live-attenuated SIV may provide important insights into the immune responses needed for protection against HIV-1. Here we investigated the development of antibodies that are functional against neutralization-resistant SIV challenge strains, and tested the hypothesis that these antibodies are associated with protection. In the absence of detectable neutralizing antibodies, Env-specific antibody-dependent cell-mediated cytotoxicity (ADCC) emerged by three weeks after inoculation with SIVDeltanef, increased progressively over time, and was proportional to SIVDeltanef replication. Persistent infection with SIVDeltanef elicited significantly higher ADCC titers than immunization with a non-persistent SIV strain that is limited to a single cycle of infection. ADCC titers were higher against viruses matched to the vaccine strain in Env, but were measurable against viruses expressing heterologous Env proteins. In two separate experiments, which took advantage of either the strain-specificity or the time-dependent maturation of immunity to overcome complete protection against SIV(mac)251 challenge, measures of ADCC activity were higher among the SIVDeltanef-inoculated macaques that remained uninfected than among those that became infected. These observations show that features of the antibody response elicited by SIVDeltanef are consistent with hallmarks of protection by live-attenuated SIV, and reveal an association between Env-specific antibodies that direct ADCC and apparent sterilizing protection by SIVDeltanef

Nucleocapsid Protein Zinc-Finger Mutants of Simian Immunodeficiency Virus Strain Mne Produce Virions That Are Replication Defectivein Vitroandin Vivo

AbstractAll retroviruses (except the spumaretroviruses) contain a nucleocapsid (NC) protein that encodes one or two copies of the Zn2+-finger sequence -Cys-X2-Cys-X4-His-X4-Cys-. This region has been shown to be essential for recognition and packaging of the genomic RNA during virion particle assembly. Additionally, this region has been shown to be involved in early infection events in a wide spectrum of retroviruses, including mammalian type C [e.g., murine leukemia virus (MuLV)], human immunodeficiency virus type 1 (HIV-1),Rous sarcomavirus, and other retroviruses. Mutations in the two Zn2+-fingers of the NC protein of simian immunodeficiency virus strain Mne [SIV(Mne)] have been generated. The resulting virions contained the normal complement of processed viral proteins with densities indistinguishable from wild-type SIV(Mne). All of the mutants had electron micrograph morphologies similar to those of immature particles observed in wild-type preparations. RNA packaging was less affected by mutations in the NC protein of SIV(Mne) than has been observed for similar mutants in the MuLV and HIV-1 systems. Nevertheless,in vitroreplication of SIV(Mne) NC mutants was impaired to levels comparable to those observed for MuLV and HIV-1 NC mutants; replication defective NC mutants are typically 105- to 106-fold less infectious than similar levels of wild-type virus. One mutant, ΔCys33–Cys36, was also found to be noninfectiousin vivowhen mutant virus was administered intravenously to a pig-tailed macaque. NC mutations can therefore be used to generate replication defective virions for candidate vaccines in the SIV macaque model for primate lentiviral diseases