Robert A. Rasmussen

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.

Molecularly Cloned SHIV-1157ipd3N4: a Highly Replication- Competent, Mucosally Transmissible R5 Simian-Human Immunodeficiency Virus Encoding HIV Clade C env

ABSTRACT Human immunodeficiency virus type 1 (HIV-1) clade C causes >50% of all HIV infections worldwide, and an estimated 90% of all transmissions occur mucosally with R5 strains. A pathogenic R5 simian-human immunodeficiency virus (SHIV) encoding HIV clade C env is highly desirable to evaluate candidate AIDS vaccines in nonhuman primates. To this end, we generated SHIV-1157i, a molecular clone from a Zambian infant isolate that carries HIV clade C env. SHIV-1157i was adapted by serial passage in five monkeys, three of which developed peripheral CD4+ T-cell depletion. After the first inoculated monkey developed AIDS at week 137 postinoculation, transfer of its infected blood to a naïve animal induced memory T-cell depletion and thrombocytopenia within 3 months in the recipient. In parallel, genomic DNA from the blood donor was amplified to generate the late proviral clone SHIV-1157ipd3. To increase the replicative capacity of SHIV-1157ipd3, an extra NF-κB binding site was engineered into its 3′ long terminal repeat, giving rise to SHIV-1157ipd3N4. This virus was exclusively R5 tropic and replicated more potently in rhesus peripheral blood mononuclear cells than SHIV-1157ipd3 in the presence of tumor necrosis factor alpha. Rhesus macaques of Indian and Chinese origin were next inoculated intrarectally with SHIV-1157ipd3N4; this virus replicated vigorously in both sets of monkeys. We conclude that SHIV-1157ipd3N4 is a highly replication-competent, mucosally transmissible R5 SHIV that represents a valuable tool to test candidate AIDS vaccines targeting HIV-1 clade C Env.

SHIV-1157i and passaged progeny viruses encoding R5 HIV-1 clade C env cause AIDS in rhesus monkeys

BackgroundInfection of nonhuman primates with simian immunodeficiency virus (SIV) or chimeric simian-human immunodeficiency virus (SHIV) strains is widely used to study lentiviral pathogenesis, antiviral immunity and the efficacy of AIDS vaccine candidates. SHIV challenges allow assessment of anti-HIV-1 envelope responses in primates. As such, SHIVs should mimic natural HIV-1 infection in humans and, to address the pandemic, encode HIV-1 Env components representing major viral subtypes worldwide.ResultsWe have developed a panel of clade C R5-tropic SHIVs based upon env of a Zambian pediatric isolate of HIV-1 clade C, the worlds most prevalent HIV-1 subtype. The parental infectious proviral clone, SHIV-1157i, was rapidly passaged through five rhesus monkeys. After AIDS developed in the first animal at week 123 post-inoculation, infected blood was infused into a sixth monkey. Virus reisolated at this late stage was still exclusively R5 tropic and mucosally transmissible. Here we describe the long-term follow-up of this initial cohort of six monkeys. Two have remained non-progressors, whereas the other four gradually progressed to AIDS within 123–270 weeks post-exposure. Two progressors succumbed to opportunistic infections, including a case of SV40 encephalitis.ConclusionThese data document the disease progression induced by the first mucosally transmissible, pathogenic R5 non-clade B SHIV and suggest that SHIV-1157i-derived viruses, including the late-stage, highly replication-competent SHIV-1157ipd3N4 previously described (Song et al., 2006), display biological characteristics that mirror those of HIV-1 clade C and support their expanded use for AIDS vaccine studies in nonhuman primates.

Neutralizing antibodies as a potential secondary protective mechanism during chronic SHIV infection in CD8+ T-cell-depleted macaques.

ObjectiveTo directly examine the role of CD8+ T cells in controlling viremia and disease during chronic, low-level primate immunodeficiency virus infection in DNA prime/protein boost-vaccinated macaques. BackgroundA cohort of macaques, vaccinated with either a DNA prime/HIV-1 gp160 boost regimen or with gp160 alone was previously protected partially from sequential challenges with non-pathogenic and pathogenic strains of chimeric simian/human immunodeficiency virus (SHIV). In this study, the effect of temporary ablation of CD8+ T cells in these animals was examined. MethodsAnimals were treated with an anti-CD8 antibody and CD8+ T-cell levels in peripheral blood, plasma viral loads, peripheral blood mononuclear cell-associated virus levels, neutralizing antibody (nAb) titers and simian immunodeficiency virus Gag-specific CD8+ T-cell numbers were followed. ResultsPlasma viremia rose sharply in direct synchrony with a rapid but transient drop in CD8+ T cells. However, although levels of cell-associated virus also rose concomitantly, peak levels were much lower than those in virus-challenged, naive animals. In addition, despite a rise of pathogenic SHIV89.6P RNA levels in three animals, CD4+ T-cell counts remained unchanged. In each of these animals, neutralizing antibody titers against the pathogenic SHIV89.6P strain were high. ConclusionsThe results indicate that CD8+ T cells play a key role in suppressing viremia in a chronically infected host. In addition, the results suggest that in the absence of CD8+ T cells, nAb may act as an effective second line of defense by limiting both the spread of infectious virus to new target cells and CD4+ T-cell loss.

Relative Transmissibility of an R5 Clade C Simian- Human Immunodeficiency Virus Across Different Mucosae in Macaques Parallels the Relative Risks of Sexual HIV-1 Transmission in Humans via Different Routes

BACKGROUND Worldwide, approximately 90% of all human immunodeficiency virus (HIV) transmissions occur mucosally; almost all involve R5 strains. Risks of sexual HIV acquisition are highest for rectal, then vaginal, and finally oral exposures. METHODS Mucosal lacerations may affect the rank order of susceptibility to HIV but cannot be assessed in humans. We measured relative virus transmissibility across intact mucosae in macaques using a single stock of SHIV-1157ipd3N4, a simian-human immunodeficiency virus encoding a primary R5 HIV clade C env (SHIV-C). RESULTS The penetrability of rhesus macaque mucosae differed significantly, with rectal challenge requiring the least virus, followed by vaginal and then oral routes (P = .031, oral vs vaginal; P < .001 rectal vs vaginal). These findings imply that intrinsic mucosal properties are responsible for the differential mucosal permeability. The latter paralleled the rank order reported for humans, with relative risk estimates within the range of epidemiological human studies. To test whether inflammation facilitates virus transmission--as predicted from human studies--we established a macaque model of localized buccal inflammation. Systemic infection occurred across inflamed but not normal buccal mucosa. CONCLUSION Our primate data recapitulate virus transmission risks observed in humans, thus establishing R5 SHIV-1157ipd3N4 in macaques as a robust model system to study cofactors involved in human mucosal HIV transmission and its prevention.

Murine and simian retrovirus models: the threshold hypothesis

By considering the dynamic relationship between retroviruses and their hosts, we have developed a unifying hypothesis to explain such disparate clinical phenomena as differential pathogenicity of a given virus in adults and neonates, transient infection with clearance of provirus-containing cells, long-term non-progression and vaccine effects of fully pathogenic viruses. The threshold hypothesis predicts that an opportunity exists during acute retroviral infection to influence the ultimate clinical outcome: if virus replication is kept below threshold by any means, including drug therapy or passive immunoprophylaxis with neutralizing antibodies, the host will prevail and win the race.

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.

Protection of neonatal macaques against experimental SHIV infection by human neutralizing monoclonal antibodies

Neonatal macaques were completely protected against oral challenge with SHIV-vpu+, a simian-human immunodeficiency virus that encodes the envelope gene of a laboratory-adapted HIV strain, by pre- and post-natal treatment with a triple combination of human neutralizing monoclonal antibodies (mAbs). The mAbs were directed either against the CD4 binding site, a glycosylation-dependent gp120 epitope, or against a linear epitope on gp41. This triple combination was highly synergistic in vitro and neutralized primary HIV completely. Subsequently, oral challenge was performed with pathogenic SHIV89.6P, an animal-passaged variant of a chimeric virus that encodes the envelope gene of the primary, dual-tropic HIV89.6. Only post-natal treatment with a similar triple mAb combination was used. One out of 4 mAb-treated infants was completely protected from infection. In the other 3 treated animals, there was a tendency towards lower peak viral RNA loads compared with untreated controls. Two out of 4 mAb-treated infants maintained normal CD4+ T-cell numbers, in contrast to all controls that had steep declines at 2 weeks post-challenge. We conclude that the triple mAb combination significantly protected the neonates, even against mucosal challenge with pathogenic SHIV89.6P. Passively administered synergistic human mAbs may play a role in preventing mother-infant transmission of HIV, both against intrapartum transmission as well as against infection through breast milk. As passive immunization is a tool to assess correlates of immune protection, we conclude that the epitopes recognized by the mAbs in our combinations are important for AIDS vaccine development. Future passive immunization studies may reveal other important conserved epitopes.

R5 clade C SHIV strains with tier 1 or 2 neutralization sensitivity: tools to dissect env evolution and to develop AIDS vaccines in primate models.

Background HIV-1 clade C (HIV-C) predominates worldwide, and anti-HIV-C vaccines are urgently needed. Neutralizing antibody (nAb) responses are considered important but have proved difficult to elicit. Although some current immunogens elicit antibodies that neutralize highly neutralization-sensitive (tier 1) HIV strains, most circulating HIVs exhibiting a less sensitive (tier 2) phenotype are not neutralized. Thus, both tier 1 and 2 viruses are needed for vaccine discovery in nonhuman primate models. Methodology/Principal Findings We constructed a tier 1 simian-human immunodeficiency virus, SHIV-1157ipEL, by inserting an “early,” recently transmitted HIV-C env into the SHIV-1157ipd3N4 backbone [1] encoding a “late” form of the same env, which had evolved in a SHIV-infected rhesus monkey (RM) with AIDS. SHIV-1157ipEL was rapidly passaged to yield SHIV-1157ipEL-p, which remained exclusively R5-tropic and had a tier 1 phenotype, in contrast to “late” SHIV-1157ipd3N4 (tier 2). After 5 weekly low-dose intrarectal exposures, SHIV-1157ipEL-p systemically infected 16 out of 17 RM with high peak viral RNA loads and depleted gut CD4+ T cells. SHIV-1157ipEL-p and SHIV-1157ipd3N4 env genes diverge mostly in V1/V2. Molecular modeling revealed a possible mechanism for the increased neutralization resistance of SHIV-1157ipd3N4 Env: V2 loops hindering access to the CD4 binding site, shown experimentally with nAb b12. Similar mutations have been linked to decreased neutralization sensitivity in HIV-C strains isolated from humans over time, indicating parallel HIV-C Env evolution in humans and RM. Conclusions/Significance SHIV-1157ipEL-p, the first tier 1 R5 clade C SHIV, and SHIV-1157ipd3N4, its tier 2 counterpart, represent biologically relevant tools for anti-HIV-C vaccine development in primates.