Amarendra Pegu

Neutralizing antibodies to HIV-1 envelope protect more effectively in vivo than those to the CD4 receptor

Neutralizing antibodies to HIV-1 envelope protect more effectively than an antibody to the CD4 cell receptor in rhesus macaques. Pushing the Envelope of HIV Protection Targeting the HIV envelope (Env) may be the best way to neutralize HIV. Pegu et al. report that broadly neutralizing antibodies to HIV Env provided more efficient protection than antibodies to the cellular receptor CD4 in rhesus macaques. Eliciting broadly neutralizing antibodies is a promising approach to preventing HIV infection. However, the best target for these antibodies has remained a matter of debate. The CD4 receptor is less variable than HIV Env, and antibodies against the CD4 receptor can potently block viral entry in vitro. Yet, when the authors compared the relative efficacy of CD4- and Env-targeting antibodies in preventing against HIV infection in macaques, they found that targeting the HIV Env may be preferable to CD4. HIV-1 infection depends on effective viral entry mediated by the interaction of its envelope (Env) glycoprotein with specific cell surface receptors. Protective antiviral antibodies generated by passive or active immunization must prevent these interactions. Because the HIV-1 Env is highly variable, attention has also focused on blocking the HIV-1 primary cell receptor CD4. We therefore analyzed the in vivo protective efficacy of three potent neutralizing monoclonal antibodies (mAbs) to HIV-1 Env compared to an antibody against the CD4 receptor. Protection was assessed after mucosal challenge of rhesus macaques with simian/HIV (SHIV). Despite its comparable or greater neutralization potency in vitro, the anti-CD4 antibody did not provide effective protection in vivo, whereas the HIV-1–specific mAbs VRC01, 10E8, and PG9, targeting the CD4 binding site, membrane-proximal, and V1V2 glycan Env regions, respectively, conferred complete protection, albeit at different relative potencies. These findings demonstrate the protective efficacy of broadly neutralizing antibodies directed to the HIV-1 Env and suggest that targeting the HIV-1 Env is preferable to the cell surface receptor CD4 for the prevention of HIV-1 transmission.

Enhanced neonatal Fc receptor function improves protection against primate SHIV infection

To protect against human immunodeficiency virus (HIV-1) infection, broadly neutralizing antibodies (bnAbs) must be active at the portals of viral entry in the gastrointestinal or cervicovaginal tracts. The localization and persistence of antibodies at these sites is influenced by the neonatal Fc receptor (FcRn), whose role in protecting against infection in vivo has not been defined. Here, we show that a bnAb with enhanced FcRn binding has increased gut mucosal tissue localization, which improves protection against lentiviral infection in non-human primates. A bnAb directed to the CD4-binding site of the HIV-1 envelope (Env) protein (denoted VRC01) was modified by site-directed mutagenesis to increase its binding affinity for FcRn. This enhanced FcRn-binding mutant bnAb, denoted VRC01-LS, displayed increased transcytosis across human FcRn-expressing cellular monolayers in vitro while retaining FcγRIIIa binding and function, including antibody-dependent cell-mediated cytotoxicity (ADCC) activity, at levels similar to VRC01 (the wild type). VRC01-LS had a threefold longer serum half-life than VRC01 in non-human primates and persisted in the rectal mucosa even when it was no longer detectable in the serum. Notably, VRC01-LS mediated protection superior to that afforded by VRC01 against intrarectal infection with simian–human immunodeficiency virus (SHIV). These findings suggest that modification of FcRn binding provides a mechanism not only to increase serum half-life but also to enhance mucosal localization that confers immune protection. Mutations that enhance FcRn function could therefore increase the potency and durability of passive immunization strategies to prevent HIV-1 infection.

Enhanced Potency of a Broadly Neutralizing HIV-1 Antibody In Vitro Improves Protection against Lentiviral Infection In Vivo

ABSTRACT Over the past 5 years, a new generation of highly potent and broadly neutralizing HIV-1 antibodies has been identified. These antibodies can protect against lentiviral infection in nonhuman primates (NHPs), suggesting that passive antibody transfer would prevent HIV-1 transmission in humans. To increase the protective efficacy of such monoclonal antibodies, we employed next-generation sequencing, computational bioinformatics, and structure-guided design to enhance the neutralization potency and breadth of VRC01, an antibody that targets the CD4 binding site of the HIV-1 envelope. One variant, VRC07-523, was 5- to 8-fold more potent than VRC01, neutralized 96% of viruses tested, and displayed minimal autoreactivity. To compare its protective efficacy to that of VRC01 in vivo, we performed a series of simian-human immunodeficiency virus (SHIV) challenge experiments in nonhuman primates and calculated the doses of VRC07-523 and VRC01 that provide 50% protection (EC50). VRC07-523 prevented infection in NHPs at a 5-fold lower concentration than VRC01. These results suggest that increased neutralization potency in vitro correlates with improved protection against infection in vivo, documenting the improved functional efficacy of VRC07-523 and its potential clinical relevance for protecting against HIV-1 infection in humans. IMPORTANCE In the absence of an effective HIV-1 vaccine, alternative strategies are needed to block HIV-1 transmission. Direct administration of HIV-1-neutralizing antibodies may be able to prevent HIV-1 infections in humans. This approach could be especially useful in individuals at high risk for contracting HIV-1 and could be used together with antiretroviral drugs to prevent infection. To optimize the chance of success, such antibodies can be modified to improve their potency, breadth, and in vivo half-life. Here, knowledge of the structure of a potent neutralizing antibody, VRC01, that targets the CD4-binding site of the HIV-1 envelope protein was used to engineer a next-generation antibody with 5- to 8-fold increased potency in vitro. When administered to nonhuman primates, this antibody conferred protection at a 5-fold lower concentration than the original antibody. Our studies demonstrate an important correlation between in vitro assays used to evaluate the therapeutic potential of antibodies and their in vivo effectiveness.

A single injection of anti-HIV-1 antibodies protects against repeated SHIV challenges

Despite the success of potent anti-retroviral drugs in controlling human immunodeficiency virus type 1 (HIV-1) infection, little progress has been made in generating an effective HIV-1 vaccine. Although passive transfer of anti-HIV-1 broadly neutralizing antibodies can protect mice or macaques against a single high-dose challenge with HIV or simian/human (SIV/HIV) chimaeric viruses (SHIVs) respectively, the long-term efficacy of a passive antibody transfer approach for HIV-1 has not been examined. Here we show, on the basis of the relatively long-term protection conferred by hepatitis A immune globulin, the efficacy of a single injection (20 mg kg−1) of four anti-HIV-1-neutralizing monoclonal antibodies (VRC01, VRC01-LS, 3BNC117, and 10-1074 (refs 9, 10, 11, 12)) in blocking repeated weekly low-dose virus challenges of the clade B SHIVAD8. Compared with control animals, which required two to six challenges (median = 3) for infection, a single broadly neutralizing antibody infusion prevented virus acquisition for up to 23 weekly challenges. This effect depended on antibody potency and half-life. The highest levels of plasma-neutralizing activity and, correspondingly, the longest protection were found in monkeys administered the more potent antibodies 3BNC117 and 10-1074 (median = 13 and 12.5 weeks, respectively). VRC01, which showed lower plasma-neutralizing activity, protected for a shorter time (median = 8 weeks). The introduction of a mutation that extends antibody half-life into the crystallizable fragment (Fc) domain of VRC01 increased median protection from 8 to 14.5 weeks. If administered to populations at high risk of HIV-1 transmission, such an immunoprophylaxis regimen could have a major impact on virus transmission.

Human Herpesvirus 8 Infects and Replicates in Primary Cultures of Activated B Lymphocytes through DC-SIGN

ABSTRACT Human herpesvirus 8 (HHV-8) is the etiological agent of Kaposis sarcoma, primary effusion lymphoma, and some forms of multicentric Castlemans disease. Although latent HHV-8 DNA can be detected in B cells from persons with these cancers, there is little information on the replication of HHV-8 in B cells. Indeed, B cells are relatively resistant to HHV-8 infection in vitro. We have recently shown that DC-SIGN, a C-type lectin first identified on dendritic cells (DC), is an entry receptor for HHV-8 on DC and macrophages. We have also demonstrated previously that B lymphocytes from peripheral blood and tonsils express DC-SIGN and that this expression increases after B-cell activation. Here we show that activated blood and tonsillar B cells can be productively infected with HHV-8, as measured by an increase in viral DNA, the expression of viral lytic and latency proteins, and the production of infectious virus. The infection of B cells with HHV-8 was blocked by the pretreatment of the cells with antibody specific for DC-SIGN or with mannan but not antibody specific for xCT, a cystine/glutamate exchange transporter that has been implicated in HHV-8 fusion to cells. The infection of B cells with HHV-8 resulted in increased expression of DC-SIGN and a decrease in the expression of CD20 and major histocompatibility complex class I. HHV-8 could also infect and replicate in B-cell lines transduced to express full-length DC-SIGN but not in B-cell lines transduced to express DC-SIGN lacking the transmembrane domain, demonstrating that the entry of HHV-8 into B cells is related to DC-SIGN-mediated endocytosis. The role of endocytosis in viral entry into activated B cells was confirmed by blocking HHV-8 infection with endocytic pathway inhibitors. Thus, the expression of DC-SIGN is essential for productive HHV-8 infection of and replication in B cells.

Safety, pharmacokinetics and neutralization of the broadly neutralizing HIV-1 human monoclonal antibody VRC01 in healthy adults

VRC‐HIVMAB060‐00‐AB (VRC01) is a broadly neutralizing HIV‐1 monoclonal antibody (mAb) isolated from the B cells of an HIV‐infected patient. It is directed against the HIV‐1 CD4 binding site and is capable of potently neutralizing the majority of diverse HIV‐1 strains. This Phase I dose‐escalation study in healthy adults was conducted at the National Institutes of Health (NIH) Clinical Center (Bethesda, MD, USA). Primary objectives were the safety, tolerability and pharmacokinetics (PK) of VRC01 intravenous (i.v.) infusion at 5, 20 or 40 mg/kg, given either once (20 mg/kg) or twice 28 days apart (all doses), and of subcutaneous (s.c.) delivery at 5 mg/kg compared to s.c. placebo given twice, 28 days apart. Cumulatively, 28 subjects received 43 VRC01 and nine received placebo administrations. There were no serious adverse events or dose‐limiting toxicities. Mean 28‐day serum trough concentrations after the first infusion were 35 and 57 μg/ml for groups infused with 20 mg/kg (n = 8) and 40 mg/kg (n = 5) doses, respectively. Mean 28‐day trough concentrations after the second infusion were 56 and 89 μg/ml for the same two doses. Over the 5–40 mg/kg i.v. dose range (n = 18), the clearance was 0·016 l/h and terminal half‐life was 15 days. After infusion VRC01 retained expected neutralizing activity in serum, and anti‐VRC01 antibody responses were not detected. The human monoclonal antibody (mAb) VRC01 was well tolerated when delivered i.v. or s.c. The mAb demonstrated expected half‐life and pharmacokinetics for a human immunoglobulin G. The safety and PK results support and inform VRC01 dosing schedules for planning HIV‐1 prevention efficacy studies.

New Member of the V1V2-Directed CAP256-VRC26 Lineage That Shows Increased Breadth and Exceptional Potency.

ABSTRACT The epitopes defined by HIV-1 broadly neutralizing antibodies (bNAbs) are valuable templates for vaccine design, and studies of the immunological development of these antibodies are providing insights for vaccination strategies. In addition, the most potent and broadly reactive of these bNAbs have potential for clinical use. We previously described a family of 12 V1V2-directed neutralizing antibodies, CAP256-VRC26, isolated from an HIV-1 clade C-infected donor at years 1, 2, and 4 of infection (N. A. Doria-Rose et al., Nature 509:55–62, 2014, http://dx.doi.org/10.1038/nature13036). Here, we report on the isolation and characterization of new members of the family mostly obtained at time points of peak serum neutralization breadth and potency. Thirteen antibodies were isolated from B cell culture, and eight were isolated using trimeric envelope probes for differential single B cell sorting. One of the new antibodies displayed a 10-fold greater neutralization potency than previously published lineage members. This antibody, CAP256-VRC26.25, neutralized 57% of diverse clade viral isolates and 70% of clade C isolates with remarkable potency. Among the viruses neutralized, the median 50% inhibitory concentration was 0.001 μg/ml. All 33 lineage members targeted a quaternary epitope focused on V2. While all known bNAbs targeting the V1V2 region interact with the N160 glycan, the CAP256-VRC26 antibodies showed an inverse correlation of neutralization potency with dependence on this glycan. Overall, our results highlight the ongoing evolution within a single antibody lineage and describe more potent and broadly neutralizing members with potential clinical utility, particularly in areas where clade C is prevalent. IMPORTANCE Studies of HIV-1 broadly neutralizing antibodies (bNAbs) provide valuable information for vaccine design, and the most potent and broadly reactive of these bNAbs have potential for clinical use. We previously described a family of V1V2-directed neutralizing antibodies from an HIV-1 clade C-infected donor. Here, we report on the isolation and characterization of new members of the family mostly obtained at time points of peak serum neutralization breadth and potency. One of the new antibodies, CAP256-VRC26.25, displayed a 10-fold greater neutralization potency than previously described lineage members. It neutralized 57% of diverse clade viral isolates and 70% of clade C isolates with remarkable potency: the median 50% inhibitory concentration was 0.001 μg/ml. Our results highlight the ongoing evolution within a single antibody lineage and describe more potent and broadly neutralizing members with potential clinical utility, particularly in areas where clade C is prevalent.

Early short-term treatment with neutralizing human monoclonal antibodies halts SHIV infection in infant macaques

Prevention of mother-to-child transmission (MTCT) of HIV remains a major objective where antenatal care is not readily accessible. We tested HIV-1–specific human neutralizing monoclonal antibodies (NmAbs) as a post-exposure therapy in an infant macaque model for intrapartum MTCT. One-month-old rhesus macaques were inoculated orally with the simian-human immunodeficiency virus SHIVSF162P3. On days 1, 4, 7 and 10 after virus exposure, we injected animals subcutaneously with NmAbs and quantified systemic distribution of NmAbs in multiple tissues within 24 h after antibody administration. Replicating virus was found in multiple tissues by day 1 in animals that were not treated. All NmAb-treated macaques were free of virus in blood and tissues at 6 months after exposure. We detected no anti-SHIV T cell responses in blood or tissues at necropsy, and no virus emerged after CD8+ T cell depletion. These results suggest that early passive immunotherapy can eliminate early viral foci and thereby prevent the establishment of viral reservoirs.

Trispecific broadly neutralizing HIV antibodies mediate potent SHIV protection in macaques

A triple threat for HIV The HIV virus continually evolves tricks to evade elimination by the host. Prevention and a cure will likely rely on broadly neutralizing antibodies that can recognize and conquer multiple viral strains or subtypes. Xu et al. engineered a single antibody molecule to recognize three highly conserved proteins needed for HIV infection (see the Perspective by Cohen and Corey). This “trispecific” antibody uses two sites (V1V2 and MPER) to bind HIV-infected cells, while the third site (CD4bs) recruits killer T lymphocytes that can eliminate the virus. When tested against >200 different HIV strains, trispecific antibodies were highly potent and broadly neutralized ∼99% of HIV viruses. This approach could potentially simplify HIV treatment regimens and improve therapy response. Science, this issue p. 85; see also p. 46 Engineered trispecific antibodies interact with three independent HIV-1 envelope determinants and prevent infection. The development of an effective AIDS vaccine has been challenging because of viral genetic diversity and the difficulty of generating broadly neutralizing antibodies (bnAbs). We engineered trispecific antibodies (Abs) that allow a single molecule to interact with three independent HIV-1 envelope determinants: the CD4 binding site, the membrane-proximal external region (MPER), and the V1V2 glycan site. Trispecific Abs exhibited higher potency and breadth than any previously described single bnAb, showed pharmacokinetics similar to those of human bnAbs, and conferred complete immunity against a mixture of simian-human immunodeficiency viruses (SHIVs) in nonhuman primates, in contrast to single bnAbs. Trispecific Abs thus constitute a platform to engage multiple therapeutic targets through a single protein, and they may be applicable for treatment of diverse diseases, including infections, cancer, and autoimmunity.

Sustained Delivery of a Broadly Neutralizing Antibody in Nonhuman Primates Confers Long-Term Protection against Simian/Human Immunodeficiency Virus Infection

ABSTRACT Pathogen-specific neutralizing antibodies protect against many viral infections and can potentially prevent human immunodeficiency virus (HIV) transmission in humans. However, neutralizing antibodies have so far only been shown to protect nonhuman primates (NHP) against lentiviral infection when given shortly before challenge. Thus, the clinical utility and feasibility of passive antibody transfer to confer long-term protection against HIV-1 are still debated. Here, we investigate the potential of a broadly neutralizing HIV-1 antibody to provide long-term protection in a NHP model of HIV-1 infection. A human antibody was simianized to avoid immune rejection and used to sustain therapeutic levels for ∼5 months. Two months after the final antibody administration, animals were completely protected against viral challenge. These findings demonstrate the feasibility and potential of long-term passive antibody for protection against HIV-1 in humans and provide a model to test antibody therapies for other diseases in NHP. IMPORTANCE Antibodies against HIV are potential drugs that may be able to prevent HIV infection in humans. However, the long-term protective capacity of antibodies against HIV has not been assessed. Here, we repetitively administered a macaque version of a human anti-HIV antibody to monkeys, after which the antibody persisted in the blood for >5 months. Moreover, the antibody could be sustained at protective levels for 108 days, conferring protection 52 days after the last dose in a monkey model of HIV infection. Thus, passive antibody transfer can provide durable protection against infection by viruses that cause AIDS in primates.