Michael Palm Basic Science, Vaccines & Prevention Project Weblog

Short Cuts To Memory

Two papers in last week’s issue of Nature report that memory CD8 T cell responses can be enhanced by drugs that alter cellular metabolism. In one of the studies, the drug rapamycin – generally used as an immune suppressant – also accelerated the development of functional memory CD8 T cells. The findings may assist efforts to develop more effective CD8 T cell vaccines and therapeutics, because one of the inconvenient aspects of T cell biology is that the differentiation process by which naïve T cells become fully functional memory T cells takes time (several weeks in mice and considerably longer in people). Furthermore, restimulation of the T cells during this differentiation process can be counterproductive, reducing the quality of the resultant memory response.

On a somewhat similar theme, the new issue of Nature Medicine includes a paper showing that manipulation of the Wnt signaling pathway can also arrest the normal T cell differentiation process, leading to the generation of memory CD8 T cells with stem cell-like self-renewal properties.

Nature 460, 41-42 (2 July 2009) | doi:10.1038/460041a; Published online 1 July 2009

News and Views

Immunology: A metabolic switch to memory

Martin Prlic & Michael J. Bevan

Abstract

Two therapeutic drugs have been found to enhance memory in immune cells called T cells, apparently by altering cellular metabolism. Are changes in T-cell metabolism the key to generating long-lived immune memory?

Nature 460, 103-107 (2 July 2009) | doi:10.1038/nature08097; Received 23 January 2009; Accepted 23 April 2009; Published online 3 June 2009

Enhancing CD8 T-cell memory by modulating fatty acid metabolism

Erika L. Pearce1, Matthew C. Walsh1, Pedro J. Cejas1, Gretchen M. Harms1, Hao Shen2, Li-San Wang1,3, Russell G. Jones4 & Yongwon Choi1

1. Department of Pathology and Laboratory Medicine,

2. Department of Microbiology,

3. Penn Center for Bioinformatics, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104, USA

4. McGill Cancer Centre, Department of Physiology, McGill University, Montreal, QC, H3G 1Y6, Canada

CD8 T cells, which have a crucial role in immunity to infection and cancer, are maintained in constant numbers, but on antigen stimulation undergo a developmental program characterized by distinct phases encompassing the expansion and then contraction of antigen-specific effector (TE) populations, followed by the persistence of long-lived memory (TM) cells Although this predictable pattern of CD8 T-cell responses is well established, the underlying cellular mechanisms regulating the transition to TM cells remain undefined. Here we show that tumour necrosis factor (TNF) receptor-associated factor 6 (TRAF6), an adaptor protein in the TNF-receptor and interleukin-1R/Toll-like receptor superfamily, regulates CD8 TM-cell development after infection by modulating fatty acid metabolism. We show that mice with a T-cell-specific deletion of TRAF6 mount robust CD8 TE-cell responses, but have a profound defect in their ability to generate TM cells that is characterized by the disappearance of antigen-specific cells in the weeks after primary immunization. Microarray analyses revealed that TRAF6-deficient CD8 T cells exhibit altered expression of genes that regulate fatty acid metabolism. Consistent with this, activated CD8 T cells lacking TRAF6 display defective AMP-activated kinase activation and mitochondrial fatty acid oxidation (FAO) in response to growth factor withdrawal. Administration of the anti-diabetic drug metformin restored FAO and CD8 TM-cell generation in the absence of TRAF6. This treatment also increased CD8 TM cells in wild-type mice, and consequently was able to considerably improve the efficacy of an experimental anti-cancer vaccine.

Nature 460, 108-112 (2 July 2009) | doi:10.1038/nature08155; Received 27 April 2009; Accepted 15 May 2009; Published online 21 June 2009

mTOR regulates memory CD8 T-cell differentiation

Koichi Araki1, Alexandra P. Turner2, Virginia Oliva Shaffer2, Shivaprakash Gangappa2, Susanne A. Keller3, Martin F. Bachmann3, Christian P. Larsen2 & Rafi Ahmed1

1. Emory Vaccine Center and Department of Microbiology and Immunology,

2. Emory Transplant Center and Department of Surgery, Emory University School of Medicine, Atlanta, Georgia 30322, USA

3. Cytos Biotechnology AG, Wagistrasse 25, 8952 Zürich-Schlieren, Switzerland

Memory CD8 T cells are a critical component of protective immunity, and inducing effective memory T-cell responses is a major goal of vaccines against chronic infections and tumours. Considerable effort has gone into designing vaccine regimens that will increase the magnitude of the memory response, but there has been minimal emphasis on developing strategies to improve the functional qualities of memory T cells. Here we show that mTOR (mammalian target of rapamycin, also known as FRAP1) is a major regulator of memory CD8 T-cell differentiation, and in contrast to what we expected, the immunosuppressive drug rapamycin has immunostimulatory effects on the generation of memory CD8 T cells. Treatment of mice with rapamycin following acute lymphocytic choriomeningitis virus infection enhanced not only the quantity but also the quality of virus-specific CD8 T cells. Similar effects were seen after immunization of mice with a vaccine based on non-replicating virus-like particles. In addition, rapamycin treatment also enhanced memory T-cell responses in non-human primates following vaccination with modified vaccinia virus Ankara. Rapamycin was effective during both the expansion and contraction phases of the T-cell response; during the expansion phase it increased the number of memory precursors, and during the contraction phase (effector to memory transition) it accelerated the memory T-cell differentiation program. Experiments using RNA interference to inhibit expression of mTOR, raptor (also known as 4932417H02Rik) or FKBP12 (also known as FKBP1A) in antigen-specific CD8 T cells showed that mTOR acts intrinsically through the mTORC1 (mTOR complex 1) pathway to regulate memory T-cell differentiation. Thus these studies identify a molecular pathway regulating memory formation and provide an effective strategy for improving the functional qualities of vaccine- or infection-induced memory T cells.

Nature Medicine 15, 731 - 732 (2009), doi:10.1038/nm0709-731

Tumor immunotherapy: making an immortal army

Brent H. Koehn1 & Stephen P. Schoenberger1

Laboratory of Cellular Immunology, La Jolla Institute for Allergy and Immunology, La Jolla, California, USA.

Abstract

Manipulation of cell renewal pathways creates T memory stem cells that can generate a sustained and targeted immune response. These findings have broad implications for vaccine development and immunotherapy (pages 808–813).

Nature Medicine 15, 808 - 813 (2009)

Published online: 14 June 2009 | doi:10.1038/nm.1982

Wnt signaling arrests effector T cell differentiation and generates CD8+ memory stem cells

Luca Gattinoni1,2, Xiao-Song Zhong1,2, Douglas C Palmer1, Yun Ji1, Christian S Hinrichs1, Zhiya Yu1, Claudia Wrzesinski1, Andrea Boni1, Lydie Cassard1, Lindsay M Garvin1, Chrystal M Paulos1, Pawel Muranski1 & Nicholas P Restifo1

1. Center for Cancer Research, National Cancer Institute, US National Institutes of Health, Bethesda, Maryland, USA.

2. These authors contributed equally to this work.

Self-renewing cell populations such as hematopoietic stem cells and memory B and T lymphocytes might be regulated by shared signaling pathways1. The Wnt–-catenin pathway is an evolutionarily conserved pathway that promotes hematopoietic stem cell self-renewal and multipotency by limiting stem cell proliferation and differentiation2, 3, but its role in the generation and maintenance of memory T cells is unknown. We found that induction of Wnt–-catenin signaling by inhibitors of glycogen sythase kinase-3 or the Wnt protein family member Wnt3a arrested CD8+ T cell development into effector cells. By blocking T cell differentiation, Wnt signaling promoted the generation of CD44lowCD62LhighSca-1highCD122highBcl-2high self-renewing multipotent CD8+ memory stem cells with proliferative and antitumor capacities exceeding those of central and effector memory T cell subsets. These findings reveal a key role for Wnt signaling in the maintenance of 'stemness' in mature memory CD8+ T cells and have major implications for the design of new vaccination strategies and adoptive immunotherapies.

Tracing HIV Reservoirs

Two new papers offer differing perspectives on the reservoirs of HIV that persist despite effective antiretroviral therapy. Nicolas Chomont and colleagues demonstrate that when memory CD4 T cells containing integrated HIV proliferate (as most memory CD4 T cells do occasionally in a process known as homeostatic self-renewal), they copy the HIV provirus along with their own genomes. When CD4 T cell numbers decline, homeostatic proliferation occurs more frequently and Chomont’s paper shows that this is associated with an increase in the number of latently infected memory CD4 T cells. The researchers describe the cells that undergo more frequent proliferation in this setting as “transitional memory” T cells. At earlier stages of infection when the CD4 T cell pool is relatively intact, the reservoir of infected memory CD4 T cells is found to be far smaller and integrated virus is primarily located in “central memory” cells that divide less frequently.

Based on these findings, the study authors suggest that anticancer drugs that interfere with memory T cell proliferation should be studied for their potential to deplete the HIV reservoir. However, given the potential toxicities associated with inhibiting T cell proliferation, the risk/benefit of such trials would need to be carefully evaluated. A more ideal therapy would be one that only targeted dividing CD4 T cells containing HIV DNA, but it is currently unclear whether such an approach is within the realm of possibility. 

The second paper - by Timothy Brennan and colleagues from Bob Siliciano’s laboratory - uses genetic analyses of HIV sequences to show that there is a reservoir of virus that seems to be coming from a cell type other than memory CD4 T cells. The study finds that in most cases, the residual virus detectable in individuals on suppressive ART is genetically distinct from the virus found in memory CD4 T cells. The authors note in their conclusion: “Numerous laboratories are actively pursuing various eradication strategies, most of which involve some aspect of targeting and purging the latent reservoir in resting memory CD4+ T cells.  If much of the residual viremia of patients undergoing HAART comes from another reservoir or compartment as suggested here, then eradication strategies will have to include ways to target and purge this additional reservoir to be successful.”

Nature Medicine

Published online: 21 June 2009 | doi:10.1038/nm.1972

HIV reservoir size and persistence are driven by T cell survival and homeostatic proliferation

Nicolas Chomont1,2,3, Mohamed El-Far1,2,3, Petronela Ancuta3, Lydie Trautmann1,2,3, Francesco A Procopio1,2,3, Bader Yassine-Diab1,2,3, Geneviève Boucher1, Mohamed-Rachid Boulassel4, Georges Ghattas5, Jason M Brenchley6, Timothy W Schacker7, Brenna J Hill8, Daniel C Douek8, Jean-Pierre Routy4,9, Elias K Haddad1,2,3,9 & Rafick-Pierre Sékaly1,2,3,9,10,11

1. Laboratoire d'Immunologie, Centre de Recherche du Centre Hospitalier de l'Université de Montréal (CR-CHUM) Saint-Luc, Montréal, Québec, Canada. 2. Laboratoire d'Immunologie, Département de Microbiologie et d'Immunologie, Université de Montréal, Québec, Canada. 3. Institute National de la Santé et de la Recherche Médicale U743, CR-CHUM, Université de Montréal, Montréal, Québec, Canada. 4. Immunodeficiency Service and Division of Hematology, Royal Victoria Hospital, McGill University Health Centre (MUHC), McGill University, Montréal, Québec, Canada. 5. Department of Gastroenterology, MUHC, Montréal, Québec, Canada. 6. Laboratory of Molecular Microbiology, National Institute of Allergy and Infectious Diseases, US National Institutes of Health, Bethesda, Maryland, USA. 7. Department of Medicine, University of Minnesota, Minneapolis, Minnesota, USA. 8. Human Immunology Section, Vaccine Research Center, National Institute of Allergy and Infectious Diseases, US National Institutes of Health, Bethesda, Maryland, USA. 9. Department of Microbiology and Immunology, McGill University, Montréal, Québec, Canada. 10. Department of Microbiology and Immunology, Oregon Health and Science University, Portland, Oregon, USA. 11. Vaccine and Gene Therapy Institute, Port-Ste Lucy, Florida, USA.

Abstract

HIV persists in a reservoir of latently infected CD4+ T cells in individuals treated with highly active antiretroviral therapy (HAART). Here we identify central memory (TCM) and transitional memory (TTM) CD4+ T cells as the major cellular reservoirs for HIV and find that viral persistence is ensured by two different mechanisms. HIV primarily persists in TCM cells in subjects showing reconstitution of the CD4+ compartment upon HAART. This reservoir is maintained through T cell survival and low-level antigen-driven proliferation and is slowly depleted with time. In contrast, proviral DNA is preferentially detected in TTM cells from aviremic individuals with low CD4+ counts and higher amounts of interleukin-7–mediated homeostatic proliferation, a mechanism that ensures the persistence of these cells. Our results suggest that viral eradication might be achieved through the combined use of strategic interventions targeting viral replication and, as in cancer, drugs that interfere with the self renewal and persistence of proliferating memory T cells.

JVI Accepts, published online ahead of print on 17 June 2009

J. Virol. doi:10.1128/JVI.02568-08

Analysis of HIV-1 Viremia and Provirus in Resting CD4+ T Cells Reveals a Novel Source of Residual Viremia in Patients on Antiretroviral Therapy

Timothy P. Brennan, John O. Woods, Ahmad R. Sedaghat, Janet D. Siliciano, Robert F. Siliciano, and Claus O. Wilke*

Department of Molecular Biology and Genetics, The Johns Hopkins University School of Medicine, Baltimore, MD 21205; Institute for Cell and Molecular Biology, The University of Texas at Austin, Austin, TX 78712; Department of Medicine, The Johns Hopkins University School of Medicine, Baltimore, MD 21205; Howard Hughes Medical Institute, Baltimore, MD 21205; Center for Computational Biology and Bioinformatics and Section of Integrative Biology, The University of Texas at Austin, Austin, TX 78712

Abstract

Highly active antiretroviral therapy (HAART) can reduce HIV-1 viremia to clinically undetectable levels. Despite this dramatic reduction, some virus is present in the blood. Additionally, a long-lived latent reservoir for HIV-1 exists in resting memory CD4+ T cells. This reservoir is believed to be a source of the residual viremia and is the focus of eradication efforts. Here, we employ two measures of population structure, analysis of molecular variance and the Slatkin-Maddison test, to demonstrate that the residual viremia is genetically distinct from proviruses in resting CD4+ T cells, but that proviruses in resting and activated CD4+ T cells belong to a single population. Residual viremia is genetically distinct from proviruses in activated CD4+ T cells, monocytes, and unfractionated peripheral blood mononuclear cells. The finding that some of the residual viremia in patients on HAART stems from an unidentified cellular source other than CD4+ T cells has implications for eradication efforts.

Illuminating Early Events in HIV Infection Using Single Genome Amplification

Two papers in the current Journal of Experimental Medicine offer unprecedented insight into the initial interactions between virus and host after HIV infection. An accompanying commentary by Zabrina Brumme and Bruce Walker eloquently articulates what these studies have achieved: “By identifying persons before seroconversion, pinpointing the transmitted virus, and assessing immune responses to that particular variant as it evolves, they provide a novel view of host and viral dynamics during the earliest stages of infection.”

In both studies the researchers use an optimized version of a technique called Single Genome Amplification (SGA), originally developed by Sarah Palmer and colleagues at the National Cancer Institute. While costly and labor-intensive, this technique allows sequencing of the HIV genome without many of the potential confounding errors that can occur with standard PCR. The researchers also used the sequences obtained by SGA to synthesize peptides for CD8 T cell response assays; this allowed detailed tracking of the impact of CD8 T cell responses on the virus genome.

The study results echo prior work from these groups suggesting that most HIV transmission events involve a single isolate; in 11 out of 12 cases SGA showed that all detected sequences were related to a single infecting virus. The remaining individual was infected with two viruses that could be unambiguously identified based on their sequences. In terms of viral evolution after infection, the researchers found that between transmission and peak viremia, diversification of HIV sequences was essentially random and showed no evidence of selection pressure from host immune responses. Subsequently, between 9-16 days later, the effects of selection became obvious, particularly effects attributable to HIV-specific CD8 T cell responses. By 32-45 days postinfection, almost the entire replicating virus population in each subject studied was replaced by viruses with mutations at two to five distinct loci in the genome, evincing selection pressure from both CD8 T cell and neutralizing antibody responses (and other unidentified sources also, perhaps innate and/or CD4 T cell immune responses).

The level of detail involved in the study also allowed the researchers to document virus escape from CD8 T cell responses earlier than has previously been reported. Mathematical modeling of the data indicated that HIV-specific CD8 T cells are more efficient at killing virus-infected cells during acute infection than prior estimates have suggested. Discussing the implications of their findings for T-cell-based vaccines, the authors state: “Modeling implied that a single T cell response was contributing as much as 15–35% of viral decline with multiple T cell responses. The implication of these observations is that vaccine-induced HIV-1–specific T cells will contribute to control of acute viremia if they are activated early in subsequent HIV-1 infection. However, because of the very rapid escape that occurs within the first few weeks of infection, T cell vaccines will need to stimulate a considerable breadth of T cell responses, clearly greater than the median of three epitopes induced by the Merck vaccine.”

Both papers are the work of researchers supported by the Center for HIV/AIDS Vaccine Immunology (CHAVI); without the substantial funding committed to this project by the National Institutes of Health, this work would not have been possible. 

Published online June 1, 2009

doi:10.1084/jem.20091094

The Journal of Experimental Medicine, Vol. 206, No. 6, 1215-1218

COMMENTARY

Tracking the culprit: HIV-1 evolution and immune selection revealed by single-genome amplification

Zabrina L. Brumme and Bruce D. Walker

Z.L. Brumme and B.D. Walker are at the Ragon Institute of MGH, MIT and Harvard, Charlestown MA 02129; Z.L. Brumme is at the Faculty of Health Sciences, Simon Fraser University, Burnaby BC V5A 1S6, Canada

ABSTRACT

Early control of HIV-1 infection is determined by a balance between the host immune response and the ability of the virus to escape this response. Studies using single-genome amplification now reveal new details about the kinetics and specificity of the CD8+ T cell response and the evolution of the virus during early HIV infection.

Published online June 1, 2009

doi:10.1084/jem.20090365

The Journal of Experimental Medicine, Vol. 206, No. 6, 1253-1272

ARTICLE

The first T cell response to transmitted/founder virus contributes to the control of acute viremia in HIV-1 infection

Nilu Goonetilleke1, Michael K.P. Liu1, Jesus F. Salazar-Gonzalez2, Guido Ferrari3, Elena Giorgi4, Vitaly V. Ganusov4, Brandon F. Keele2, Gerald H. Learn2, Emma L. Turnbull5, Maria G. Salazar2, Kent J. Weinhold3, Stephen Moore1, CHAVI Clinical Core B, Norman Letvin6, Barton F. Haynes3, Myron S. Cohen7, Peter Hraber4, Tanmoy Bhattacharya4,8, Persephone Borrow5, Alan S. Perelson4, Beatrice H. Hahn2, George M. Shaw2, Bette T. Korber4,8, and Andrew J. McMichael1

1 Medical Research Council Human Immunology Unit, Weatherall Institute of Molecular Medicine, Oxford University, Oxford OX3 9DS, England, UK

2 Department of Microbiology, University of Alabama at Birmingham, Birmingham, AL 35294

3 Duke University Medical Research, Duke University, Durham, NC 27710

4 Los Alamos National Laboratory, Theoretical Division, Los Alamos, NM 87545

5 The Jenner Institute, Oxford University, Compton RG20 7NN, England, UK

6 BIDMC, Harvard University, Boston, MA 02115

7 HIV Prevention Trials Unit, University of North Carolina, Chapel Hill, NC 27599

8 The Santa Fe Institute, Santa Fe, NM 87501

Identification of the transmitted/founder virus makes possible, for the first time, a genome-wide analysis of host immune responses against the infecting HIV-1 proteome. A complete dissection was made of the primary HIV-1–specific T cell response induced in three acutely infected patients. Cellular assays, together with new algorithms which identify sites of positive selection in the virus genome, showed that primary HIV-1–specific T cells rapidly select escape mutations concurrent with falling virus load in acute infection. Kinetic analysis and mathematical modeling of virus immune escape showed that the contribution of CD8 T cell–mediated killing of productively infected cells was earlier and much greater than previously recognized and that it contributed to the initial decline of plasma virus in acute infection. After virus escape, these first T cell responses often rapidly waned, leaving or being succeeded by T cell responses to epitopes which escaped more slowly or were invariant. These latter responses are likely to be important in maintaining the already established virus set point. In addition to mutations selected by T cells, there were other selected regions that accrued mutations more gradually but were not associated with a T cell response. These included clusters of mutations in envelope that were targeted by NAbs, a few isolated sites that reverted to the consensus sequence, and bystander mutations in linkage with T cell–driven escape.

Published online June 1, 2009

doi:10.1084/jem.20090378

The Journal of Experimental Medicine, Vol. 206, No. 6, 1273-1289

ARTICLE

Genetic identity, biological phenotype, and evolutionary pathways of transmitted/founder viruses in acute and early HIV-1 infection

Jesus F. Salazar-Gonzalez1, Maria G. Salazar1, Brandon F. Keele1, Gerald H. Learn1, Elena E. Giorgi2,3, Hui Li1, Julie M. Decker1, Shuyi Wang1, Joshua Baalwa1, Matthias H. Kraus1, Nicholas F. Parrish1, Katharina S. Shaw1, M. Brad Guffey1, Katharine J. Bar1, Katie L. Davis1, Christina Ochsenbauer-Jambor1, John C. Kappes1, Michael S. Saag1, Myron S. Cohen4, Joseph Mulenga5, Cynthia A. Derdeyn6, Susan Allen6, Eric Hunter6, Martin Markowitz7,8, Peter Hraber2, Alan S. Perelson2, Tanmoy Bhattacharya2,9, Barton F. Haynes10, Bette T. Korber2,9, Beatrice H. Hahn1, and George M. Shaw1

1 University of Alabama at Birmingham, Birmingham, AL 35294

2 Los Alamos National Laboratory, Los Alamos, NM 87545

3 University of Massachusetts, Amherst, MA 01002

4 The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599

5 Zambia-Emory HIV Research Project, Lusaka, Zambia

6 Emory University, Atlanta, GA 30329

7 Aaron Diamond AIDS Research Center, New York, NY 10016

8 The Rockefeller University, New York, NY 10065

9 Santa Fe Institute, Santa Fe, NM 87501

10 Duke University Medical Center, Durham, NC 27710

Identification of full-length transmitted HIV-1 genomes could be instrumental in HIV-1 pathogenesis, microbicide, and vaccine research by enabling the direct analysis of those viruses actually responsible for productive clinical infection. We show in 12 acutely infected subjects (9 clade B and 3 clade C) that complete HIV-1 genomes of transmitted/founder viruses can be inferred by single genome amplification and sequencing of plasma virion RNA. This allowed for the molecular cloning and biological analysis of transmitted/founder viruses and a comprehensive genome-wide assessment of the genetic imprint left on the evolving virus quasispecies by a composite of host selection pressures. Transmitted viruses encoded intact canonical genes (gag-pol-vif-vpr-tat-rev-vpu-env-nef) and replicated efficiently in primary human CD4+ T lymphocytes but much less so in monocyte-derived macrophages. Transmitted viruses were CD4 and CCR5 tropic and demonstrated concealment of coreceptor binding surfaces of the envelope bridging sheet and variable loop 3. 2 mo after infection, transmitted/founder viruses in three subjects were nearly completely replaced by viruses differing at two to five highly selected genomic loci; by 12–20 mo, viruses exhibited concentrated mutations at 17–34 discrete locations. These findings reveal viral properties associated with mucosal HIV-1 transmission and a limited set of rapidly evolving adaptive mutations driven primarily, but not exclusively, by early cytotoxic T cell responses.

Natural Neutralizing Antibodies

A number of recent studies have reported the detection of neutralizing antibodies in individuals with chronic HIV infection. In some rare cases, antibody responses capable of neutralizing a broad array of diverse HIV isolates have been documented. In a perspective piece just published by Nature Medicine, Leonidas Stamatatos and colleagues review these findings and suggest that they represent good news for the vaccine field. In particular, they note the data argue that B cells can make antibodies capable of neutralizing HIV; some scientists have been concerned that the antibody structure required to inhibit HIV cannot be created by the human immune system. Based on the new findings, the researchers recommend a series of steps for pursuing structure-based HIV vaccine design:

 

Figure 1 - Flow chart of structure-based HIV-1 vaccine design.

CREDIT:

Nature Medicine

Published online: 14 June 2009 | doi:10.1038/nm.1949

Neutralizing antibodies generated during natural HIV-1 infection: good news for an HIV-1 vaccine?

Leonidas Stamatatos1,2, Lynn Morris3,4, Dennis R Burton5 & John R Mascola6

1. Seattle Biomedical Research Institute, Seattle, Washington, USA.

2. Department of Global Health, University of Washington, Seattle, Washington, USA.

3. AIDS Virus Research Unit, National Institute for Communicable Diseases, Johannesburg, South Africa.

4. Centre for the AIDS Programme of Research in South Africa, Johannesburg, South Africa.

5. Department of Immunology and Microbial Science and the International AIDS Vaccine Initiative Neutralizing Antibody Center, The Scripps Research Institute, La Jolla, California, USA.

6. Vaccine Research Center, National Institute of Allergy and Infectious Diseases, US National Institutes of Health, Bethesda, Maryland, USA.

Abstract

Most existing viral vaccines generate antibodies that either block initial infection or help eradicate the virus before it can cause disease. For HIV-1, obstacles to eliciting protective neutralizing antibodies (NAbs) have often seemed insurmountable. The target of HIV-specific NAbs, the viral envelope glycoprotein (Env), is highly variable in amino acid sequence and glycosylation pattern. Conserved elements of HIV-1 Env seem to be poorly immunogenic, and previous attempts to generate broadly reactive NAbs by vaccination have proven ineffective. However, recent studies show that antibodies in the sera of some HIV-1–infected individuals can neutralize diverse HIV-1 isolates. Detailed analyses of these sera provide new insights into the viral epitopes targeted by broadly reactive NAbs. The findings discussed here suggest that the natural NAb response to HIV-1 can inform future vaccine design. A concerted effort of structure-based vaccine design will help guide the development of improved antibody-based vaccines for HIV-1.

Killer Helpers

CD4 T cells are typically portrayed as playing a supportive role in the generation and maintenance of immune responses, as their alternative name – helper cells – suggests. But over the past decade, studies have shown that there are a number of settings in which antigen-specific CD4 T cells exert direct cytotoxic activity (a capacity that was once believed to be an artifact of prolonged in vitro culture).

Two new papers describe the presence of cytotoxic virus-specific CD4 T cells in SIV and HIV infection, respectively. The first study, led by Jonah Sacha, obtained Gag- and Nef-specific CD4 T cells from a group of elite controller macaques infected with SIVmac239. The researchers had noticed that, after an experiment in which CD8 T cells were depleted, the animals experienced a rebound in viral load that was subsequently brought back down to undetectable levels. This regaining of control over viral replication was associated with an expansion of their Gag- and Nef-specific CD4 T cell responses, an observation that prompted the current study.

In Sacha’s in vitro analyses, cytotoxic Gag- and Nef-specific CD4 T cells were unable to recognize SIV-infected CD4 T cells, but efficiently recognized and killed infected macrophages. Recognition occurred very rapidly, within 1-2 hours, with activity peaking between 6 and 24 hours. The cytotoxic CD4 T cells consistently eliminated 30-40% of SIV-infected macrophages in culture. Sacha and colleagues conclude by stating “on the basis of the data presented here, we speculate that Gag- and Nef-specific CD4􏰀 T cells may play a more important role in the antiretroviral immune response than previously appreciated.”

The second paper, by Nan Zheng and colleagues from Kumamoto University, identifies cytotoxic Nef-specific CD4 T cells in 4/9 individuals analyzed. In this study, the Nef-specific CD4 T cells are reported to kill both HIV-infected macrophages and CD4 T cells, but the activity against macrophages is shown to be superior. 

Taken together, the findings suggest that it might be worth investigating whether cytotoxic virus-specific CD4 T cells can be induced by vaccination. Perhaps of note, one previous report from Ron Desrosiers laboratory has suggested that effector CD4 T cells with a cytotoxic phenotype (expressing perforin and the degranulation marker CD107) contribute to the protection obtained with live-attenuated SIV vaccines.

The PNAS paper is Open Access, click the title link for the full PDF. 

PNAS Published online before print May 28, 2009, doi: 10.1073/pnas.0813106106

Gag- and Nef-specific CD4+ T cells recognize and inhibit SIV replication in infected macrophages early after infection

Jonah B. Sacha a,1,2, Juan P. Giraldo-Vela a,1, Matthew B. Buechler a, Mauricio A. Martins a, Nicholas J. Maness a, Chungwon Chung a, Lyle T. Wallace a, Enrique J. León a, Thomas C. Friedrich b, Nancy A. Wilson a, Atsunobu Hiraoka  and David I. Watkins a

a Departments of aPathology and Laboratory Medicine andb  Pathobiological Sciences, University of Wisconsin, Madison, WI 53715; and c Osaka Dental University, Osaka 540-0008, Japan

1J.B.S. and J.P.G.-V. contributed equally to this work.

Abstract

The precise immunological role played by CD4+ T cells in retroviral infections is poorly defined. Here, we describe a new function of these cells, the elimination of retrovirus-infected macrophages. After experimental CD8+ cell depletion, elite controlling macaques with set-point viral loads ≤500 viral RNA copies/mL mounted robust Gag- and Nef-specific CD4+ T cell responses during reestablishment of control with ≥54% of all virus-specific CD4+ T cells targeting these 2 proteins. Ex vivo, these simian immunodeficiency virus (SIV)-specific CD4+ T cells neither recognized nor suppressed viral replication in SIV-infected CD4+ T cells. In contrast, they recognized SIV-infected macrophages as early as 2 h postinfection because of presentation of epitopes derived from virion-associated Gag and Nef proteins. Furthermore, virus-specific CD4+ T cells displayed direct effector function and eliminated SIV-infected macrophages. These results suggest that retrovirus-specific CD4+ T cells may contribute directly to elite control by inhibiting viral replication in macrophages.

JVI Accepts, published online ahead of print on 20 May 2009

J. Virol. doi:10.1128/JVI.00513-09

Strong ability of Nef-specific CD4+ cytotoxic T cells to suppress HIV-1 replication in HIV-1-infected CD4+ T cells and macrophages

Nan Zheng, Mamoru Fujiwara, Takamasa Ueno, Shinichi Oka, and Masafumi Takiguchi

Division of Viral Immunology and Division of Infectious Disease, Center for AIDS Research, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811; and AIDS Clinical Center, International Medical Center of Japan, 1-21-1, Toyama, Shinjuku, Tokyo 162-8655, Japan

Abstract

A restricted number of studies have shown that HIV-1-specific cytotoxic CD4+ T cells are present in HIV-1-infected individuals. However, the roles of this type of CD4+ T cell in the immune responses against an HIV-1 infection remain unclear. In this study, we identified novel Nef epitope-specific HLA-DRB1*0803-restricted cytotoxic CD4+ T cells. The CD4+ T cell clones specific for Nef187-203 showed strong IFN-production after having been stimulated with autologous B-LCLs infected with Nef recombinant vaccinia virus or pulsed with heat-inactivated virus particles, indicating the presentation of the epitope antigen through both exogenous and endogenous MHC class II processing pathways. Nef187-203-specific CD4+ T cell clones exhibited strong cytotoxic activity against both HIV-1-infected macrophages and CD4+ T cells from an HLA-DRB1*0803+ donor. In addition, these Nef-specific cytotoxic CD4+ T cell clones exhibited strong ability to suppress HIV-1 replication in both macrophages and in CD4+ T cells in vitro. Nef187-203-specific cytotoxic CD4+ T cells were detected in cultures of peptide-stimulated PBMC and in ex vivo PBMC from 40% and 20% of DRB1*0803+ donors, respectively. These results suggest that HIV-1-specific CD4+ T cells may directly control HIV-1-infection in vivo by suppressing virus replication in HIV-1 natural host cells.

Maximum Suppression: ART Intensification Does Not Reduce Residual Viral Load

One of the most controversial questions in the field of HIV research is whether current antiretroviral therapy (ART) combinations maximally suppress viral replication. New technologies have allowed researchers to detect down to just 1 copy of HIV RNA per mL of blood and, even when the viral load is undetectable on commercially available tests (which typically can only detect 50 copies or more), most people on ART have a few copies of HIV RNA detectable in their samples. These HIV RNA copies could either be the product of ongoing rounds of viral replication (in which infected cells release new viruses that go on to infect other cells), or alternatively they could be produced by long-lived chronically infected cells. In the latter scenario, ART would prevent the newly-produced virus from being able to infect any other cells, but the drugs would not be able to eliminate the chronically infected cell.

Over the past few years, scientists have debated – often quite heatedly – which of these possibilities is true. Recent evidence has generally favored the view that, in most cases, ART is maximally suppressing HIV replication; for example, a study of viral evolution in people undergoing intermittent ART interruption found no evidence of ongoing viral evolution during the periods when participants were on therapy.

A study just published in PNAS tackles the question another way, by investigating whether intensifying ART by adding new drugs has an effect on residual viral load. Out of 15 total participants, only 9 consistently showed HIV RNA levels above 1 copy/mL prior to ART intensification (median 3 copies/mL). The highest level detected was around 30 copies/mL. The researchers found that ART intensification had no effect on these residual viral levels, indicating a lack of ongoing HIV replication. The results add to the evidence that low-level HIV RNA detectable in people on ART does not derive from ongoing viral replication, but rather a stable reservoir of infected cells. The major implication is that, in order to cure HIV infection, new strategies are needed to identify and eliminate this reservoir. PNAS has designated the paper "Open Access," click on the title link for the PDF. 

PNAS

Published online before print May 22, 2009, doi: 10.1073/pnas.0903107106

Treatment intensification does not reduce residual HIV-1 viremia in patients on highly active antiretroviral therapy

J. B. Dinoso a,b,1, S. Y. Kim a,1, A. M. Wiegand c, S. E. Palmer c,2, S. J. Gange d, L. Cranmer a, A. O'Shea e, M. Callender a, A. Spivak a, T. Brennan a, M. F. Kearney c, M. A. Proschan f, J. M. Mican g, C. A. Rehm g, J. M. Coffin c,h,3, J. W. Mellors i, R. F. Siliciano a,j and F. Maldarelli c,3

Departments of a Medicine and b Pharmacology and Molecular Sciences, The Johns Hopkins University School of Medicine, Baltimore, MD 21205;

c HIV Drug Resistance Program, National Cancer Institute-Frederick, National Institutes of Health, Frederick, MD 21702;

d Department of Epidemiology, The Johns Hopkins University Bloomberg School of Public Health, Baltimore, MD 21205;

e Critical Care Medicine Department, Warren Grant Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892;

f Biostatistics Research Branch and g Division of Clinical Research, National Institute of Allergy and Infectious Diseases, Bethesda, MD 20892;

h Department of Molecular Biology and Microbiology, Tufts University, Boston, MA 02111;

i Department of Infectious Diseases, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261; and

j Howard Hughes Medical Institute, The Johns Hopkins University School of Medicine, Baltimore, MD 21205

1 J.B.D. and S.Y.K. contributed equally to this work.

Abstract

In HIV-1-infected individuals on currently recommended antiretroviral therapy (ART), viremia is reduced to <50 copies of HIV-1 RNA per milliliter, but low-level residual viremia appears to persist over the lifetimes of most infected individuals. There is controversy over whether the residual viremia results from ongoing cycles of viral replication. To address this question, we conducted 2 prospective studies to assess the effect of ART intensification with an additional potent drug on residual viremia in 9 HIV-1-infected individuals on successful ART. By using an HIV-1 RNA assay with single-copy sensitivity, we found that levels of viremia were not reduced by ART intensification with any of 3 different antiretroviral drugs (efavirenz, lopinavir/ritonavir, or atazanavir/ritonavir). The lack of response was not associated with the presence of drug-resistant virus or suboptimal drug concentrations. Our results suggest that residual viremia is not the product of ongoing, complete cycles of viral replication, but rather of virus output from stable reservoirs of infection.

Interrogating the Gut's Contribution to Peak Viral Load

A new study from Miles Davenport and colleagues, just published online in the Journal of Virology, uses repeated sampling of SIV-infected monkeys to show that, in the majority of cases, gut CD4 T cell loss happens so quickly that these cells cannot contribute substantially to the high levels of viral load that occur during acute SIV infection. The authors note that prior studies did not sample animals with sufficient frequency to detect that loss of gut CD4 T cells typically precedes peak viral load by 4-5 days. In contrast, the researchers observed that peak CD4 T cell loss from lymphoid tissues, bronchoalveolar lavage (BaL) and peripheral blood coincided with peak viral load. 

JVI Accepts, published online ahead of print on 20 May 2009

J. Virol. doi:10.1128/JVI.00552-09

Is gut the major source of virus in early SIV infection?

Matthew D. H. Lay, Janka Petravic, Shari N. Gordon, Jessica Engram, Guido Silvestri, and Miles P Davenport*

Complex Systems in Biology Group, Centre for Vascular Research, University of New South Wales, Kensington NSW 2052, Australia; Departments of Pathology & Laboratory Medicine, and Microbiology, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania; Yerkes National Primate Research Center, Emory University, Atlanta, GA, USA

The acute phases of HIV and SIV infection are characterized by rapid and profound depletion of CD4+ T cells from the gut of infected individuals. The large number of CD4+ T cells in the gut (a large fraction of which is activated and expresses the HIV/SIV co-receptor CCR5), the high level of infection of these cells, and the temporal coincidence of this CD4+ T cell depletion with the peak of virus in plasma in acute infection suggest that the intestinal mucosa may be the major source of virus driving the peak viral load. Here we use data on CD4+ T cell proportions in the lamina propria of the rectum of SIV infected rhesus macaques (that progress to AIDS) and sooty mangabeys (that do not progress) to show that, in both species, the depletion of CD4+ T cells from this mucosal site and its maximum loss rate are often observed several days before the peak in viral load, with few CD4+ T cells remaining in the rectum by the time of peak viral load. By contrast, the maximum loss rate of CD4+ T cells from broncho-alveolar lavage and lymph node coincides with the peak in virus. Analysis of the kinetics of depletion suggests that, in both rhesus macaques and sooty mangabeys, CD4+ T cells in the intestinal mucosa are a highly susceptible population for infection but not a major source of plasma virus in acute SIV infection.

New Data on the Association Between Lymphopenia & Microbial Translocation

Several prior blog posts have covered the issue of microbial translocation (the leaking of normally friendly gut bacteria into systemic circulation) in people with HIV. There is now a considerable body of evidence that microbial translocation occurs in chronic HIV infection, but the mechanism remains unclear. The original paper by Jason Brenchley and colleagues suggested that gut CD4 T cell depletion by HIV during acute infection was to blame, but there was no evidence of microbial translocation until later in the progression of disease. The standard measure of microbial translocation is plasma levels of lipopolysaccharide (LPS), and Brenchley et al suggested that anti-LPS antibodies present in early infection effectively mopped up systemic LPS and only when these antibodies waned did levels become detectable.

However, a subsequent mouse study raised the possibility that there might be a connection between loss of naïve CD4 T cells – which happens slowly but progressively as HIV infection advances  - and microbial translocation. It turns out that this has been a longstanding observation in the mouse literature, with a 1980 paper documenting microbial translocation in mice bred to lack a thymus, which could be ameliorated when the mice were given thymic grafts.  

In the new issue of the Journal of Infectious Diseases, Philip Lee from Irini Sereti’s lab at NIAID sheds further light on the issue with a study of microbial translocation in individuals with idiopathic CD4 T cell lymphopenia (ICL), which is characterized by low CD4 and CD8 T cell levels in the absence of a known cause. In the early years of the HIV epidemic, cases of ICL were briefly a source of controversy due to claims that they represented AIDS without HIV. Studies have since made clear the significant differences between ICL and AIDS: there is no CD8 T cell activation in ICL, whereas the level of CD8 T cell activation is the single strongest predictor of disease progression in HIV infection. CD8 T cell numbers also decline in ICL but increase in HIV until disease becomes very advanced. People with HIV also accumulate both CD4 and CD8 T cell responses to the protein products of all nine HIV genes, such that 20% or more of all memory CD8 T cells can be shown to be HIV-specific by interferon gamma ELISPOT in individuals with an AIDS diagnosis (HIV-specific immune responses are of course entirely absent in ICL).

Lee and colleagues conducted their study to explore whether ICL is associated with microbial translocation. Eleven individuals with ICL were enrolled, along with ten HIV-infected individuals not on ART and 8 HIV-negative healthy controls. Confirming prior results, CD8 counts were significantly elevated in people with HIV compared to controls, while CD8 counts in participants with ICL were significantly lower than controls. CD8 T cell activation was also significantly elevated in HIV infection, but not in controls or individuals with ICL.

The novel and intriguing finding of the study was that plasma LPS levels were significantly elevated – to a similar extent - in both HIV infection and ICL compared to controls. In people with HIV there was a significant association between LPS levels and the percentage of naïve CD4 T cells (r=-0.68; p=.035) but this relationship was not seen in ICL. The researchers also found a significant correlation between the percentage of proliferating CD4 T cells (as measured by Ki67 expression) and LPS levels in ICL (r=-0.88;p=.003), but not in participants with HIV.

The discussion section of the paper cites murine studies indicating that gut bacteria antigens can contribute to lymphopenia-induce CD4 T cell proliferation, and notes that “in individuals with CD4 lymphopenia, including HIV-infected individuals, translocation of microbial products may play an integral role in lymphopenia-induced proliferation and chronic immune activation, or it may simply be the result of CD4 T cell lymphopenia either in the gut or in the lymphoid tissue.”

The finding that the presence of elevated LPS levels is not associated with CD8 T cell activation in ICL is also highlighted. In the initial study of microbial translocation in HIV infection by Jason Brenchley (who is also a co-author on this paper), there was a statistically significant correlation between LPS levels and CD8 T cell activation and systemic LPS was suggested to be playing a causative role. The current study seems to run counter to that, possibility, but, as the authors state, “intermediary steps of the innate immune response may be vastly different in HIV-infected patients.” In other words, it is plausible that the altered immune environment in untreated HIV infection (including factors such as increased antigen-presentation and antigen-presenting cell turnover) could impact the interaction between LPS and CD8 T cells. Alternatively, the researchers acknowledge the possibility that other antigens may be the primary stimulus of CD8 T cell activation in the setting of HIV infection.

Additional studies of the impact of ICL on CD4 T cell numbers in gut and lymphoid tissue should help discern the commonalities and differences in the pathogenesis of microbial translocation in ICL and HIV. 

The Journal of Infectious Diseases 2009;199:1664–1670

DOI: 10.1086/598953

MAJOR ARTICLE

Evidence for Translocation of Microbial Products in Patients with Idiopathic CD4 Lymphocytopenia

Philip I. Lee,1  Emily J. Ciccone,1  Sarah W. Read,2  Ava Asher,3  Robert Pitts,1  Daniel C. Douek,3  Jason M. Brenchley,4 and Irini Sereti1

1Laboratory of Immunoregulation, Clinical and Molecular Retrovirology Section, 2Division of AIDS, 3Human Immunology Section, Vaccine Research Center, and 4Laboratory of Molecular Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland

Translocation of microbial products has been described in chronic human immunodeficiency virus (HIV) infection and correlates with activation of the immune system. We investigated the potential translocation of microbial products in idiopathic CD4 lymphocytopenia (ICL), a rare disorder characterized by low CD4 T cell counts in the absence of HIV infection. Plasma lipopolysaccharide (LPS) levels and T cell activation were measured in a cross‐sectional cohort study of patients with ICL and HIV infection and healthy control subjects. Increases in CD4 T cell proliferation but not CD8 T cell proliferation were observed in patients with ICL. LPS levels were significantly elevated both in patients with ICL and in patients with HIV infection, and they were strongly correlated with the proportion of proliferating CD4 T cells in the cohort of patients with ICL (r=0.88;p=.003). The proportions of T helper (Th) 17 and Th1 CD4 cells in peripheral blood were similar between patients with ICL, patients with HIV infection, and control subjects. These findings suggest a potential association of translocation of microbial products with perturbed CD4 T cell homeostasis in individuals with CD4 lymphopenic states other than HIV infection.

Genetically Engineered Immunity

Today (May 18) is HIV Vaccine Awareness Day and, with an eye for a PR opportunity, Nature Medicine has just published a study of a novel approach to immunization against HIV infection. Philip Johnson and colleagues describe their use of an adeno-associated virus (AAV) vector to persistently express antibody-like “immunoadhesins” that can inhibit a particular SIV variant called SIVmac316. They report that delivery of the AAV vector to rhesus macaques produced sustained circulating levels of immunoadhesins that protected 6/9 animals from a subsequent intravenous SIVmac316 challenge.

The paper has garnered a fair amount of media coverage today. While the approach is novel and the results are somewhat encouraging, there are some important caveats to be borne in mind. Firstly, it is as yet unclear whether AAV can be engineered to produce similar sustained protein expression in humans. While it’s not mentioned in the paper, a previous AAV vector - developed by Philip Johnson and manufactured by Targeted Genetics - was designed to encode HIV proteins with the goal of triggering the development of HIV-specific T cell responses. This AAV vector proved dismally immunogenic in phase I human trials, inducing detectable HIV-specific T cell responses in fewer than 1 in 5 participants.

Although this new study obviously represents a different use of the vector, the lack of immunogenicity in prior trials suggests that it was not efficient at producing proteins in the human body. The results echo the disappointments of other research groups who have attempted to use AAV as a gene transfer vehicle.  As reported on the blog previously, the explanation may be that many people have pre-existing CD8 T cell responses against the AAV capsid, which can cross-react with capsids from different AAV serotypes. Killing of AAV-infected cells by these CD8 T cells would limit the ability of the vector to encode its protein payload. Macaques (and other animals used in preclinical studies) do not have pre-existing AAV-specific CD8 T cell responses.

The other major caveat regarding the approach is the potential for the induction of immune responses against the antibody or immunoadhesin being produced by the vector. The three immunized macaques that became SIV-infected in Johnson’s study all showed evidence of antibody responses against the immunoadhesin. However, it is perhaps encouraging to note that a human trial that administered a combination of broadly neutralizing monoclonal antibodies (2G12, 4E10 and 2F5) against HIV did not detect the generation of an immune response against the antibodies, at least in the setting of extant HIV infection (to the best of my knowledge, the antibodies have not been administered to HIV-negative volunteers,).

The overall idea of genetically engineered immunity is certainly novel, and timely given the struggles involved in persuading the adaptive immune system to make a response that protects against HIV infection. An excellent description of the approaches under study, including Johnson’s, can be found in an article by Andreas von Bubnoff from the July-August 2008 issue of IAVI Report. What remains unclear, despite these new data, is whether AAV can work as a vehicle for delivering antibodies in humans.

Nature Medicine

Published online: 17 May 2009 | doi:10.1038/nm.1967

Vector-mediated gene transfer engenders long-lived neutralizing activity and protection against SIV infection in monkeys

Philip R Johnson1, Bruce C Schnepp1, Jianchao Zhang2, Mary J Connell1, Sean M Greene1, Eloisa Yuste3, Ronald C Desrosiers3 & K Reed Clark2

1. The Children's Hospital of Philadelphia and the University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, USA.

2. The Research Institute at Nationwide Children's Hospital, Columbus, Ohio, USA.

3. New England Primate Research Center and Harvard Medical School, Southborough, Massachusetts, USA.

Abstract

The key to an effective HIV vaccine is development of an immunogen that elicits persisting antibodies with broad neutralizing activity against field strains of the virus. Unfortunately, very little progress has been made in finding or designing such immunogens. Using the simian immunodeficiency virus (SIV) model, we have taken a markedly different approach: delivery to muscle of an adeno-associated virus gene transfer vector expressing antibodies or antibody-like immunoadhesins having predetermined SIV specificity. With this approach, SIV-specific molecules are endogenously synthesized in myofibers and passively distributed to the circulatory system. Using such an approach in monkeys, we have now generated long-lasting neutralizing activity in serum and have observed complete protection against intravenous challenge with virulent SIV. In essence, this strategy bypasses the adaptive immune system and holds considerable promise as a unique approach to an effective HIV vaccine.

2G12: The Sweetest Broadly Neutralizing Antibody

A number of monoclonal antibodies with broad neutralizing activity against HIV have been identified, and given names suggestive of characters from a futuristic George Lucas movie: 2F5, b12, 4E10 and 2G12. Despite their broad activity, macaque experiments in which the antibodies are infused prior to a virus challenge have shown that, in most cases, dauntingly high antibody titers are required to obtain sterilizing protection. Even if a means to induce similar antibodies via vaccination can be found, it is unlikely that such high antibody levels could be achieved, or sustained. But all may not be lost, because one study – conducted by John Mascola and colleagues many years ago - has hinted that the antibody 2G12 may be an exception to this rule. 2G12 is unique among the broadly neutralizing monoclonals because it targets the glycan structures on HIV’s envelope that usually function to shield the virus from antibody-mediated neutralization (a glycan is a large carbohydrate molecule composed of many smaller sugar molecules linked together).

Ann Hessell and colleagues from Dennis Burton’s research group decided to follow up on Mascola’s lead, and have now published their results in PLoS Pathogens. They note that the original experiment used the much-maligned X4-using SHIV89.6P as a challenge virus, and the goal of the new study was to test the concentration of 2G12 required to protect against SHIVSF162p3, which has an R5-using HIV envelope that is only mildly susceptible to 2G12 in vitro. Five macaques received an infusion of 2G12 prior to intravaginal challenge, while two controls received control antibody and two additional controls were untreated. All four controls became infected, whereas three out of five 2G12 recipients were completely protected from infection. One of the remaining macacques had a delay in the appearance of virus and diminished replication, while the other displayed a viral load peak and set point that was indistinguishable from the control group. The antibody titer required to achieve this outcome was 1:1, substantially lower than the level of around 1:100 previously reported for b12.

In discussing the results, Hessell et al evaluate several potential explanations for the apparent ability of 2G12 to protect at lower titers. Enhanced antibody trafficking to the site of exposure and superior antibody-dependent cellular cytotoxicity (ADCC) did not appear to account for the outcome. The researchers currently favor a role for the enhanced neutralization kinetics that are observed in vitro with 2G12 compared to other antibodies. They also note that 2G12 has the ability to block the interaction between gp120 and the dendritic cell molecule DC-SIGN, which could also contribute to superior protective efficacy. Additional experiments are needed to fully explore these possibilities, but as the authors state: “the results are provocative in suggesting the glycan shield as a potentially favorable HIV vaccine target.”

PLoS Pathog 5(5): e1000433. doi:10.1371/journal.ppat.1000433

Broadly Neutralizing Human Anti-HIV Antibody 2G12 Is Effective in Protection against Mucosal SHIV Challenge Even at Low Serum Neutralizing Titers

Ann J. Hessell1, Eva G. Rakasz2, Pascal Poignard1,3, Lars Hangartner1,4, Gary Landucci5, Donald N. Forthal5, Wayne C. Koff3, David I. Watkins2, Dennis R. Burton1*

1 Department of Immunology and Microbial Science, and IAVI Neutralizing Antibody Center, The Scripps Research Institute, La Jolla, California, United States of America, 2 Department of Pathology and Laboratory Medicine, University of Wisconsin-Madison, Madison, Wisconsin, United States of America, 3 International AIDS Vaccine Initiative (IAVI), New York, New York, United States of America, 4 Institute of Medical Virology, University of Zürich, Zürich, Switzerland, 5 Division of Infectious Diseases, Department of Medicine, UC Irvine School of Medicine, University of California Irvine, Irvine, California, United States of America

Developing an immunogen that elicits broadly neutralizing antibodies (bNAbs) is an elusive but important goal of HIV vaccine research, especially after the recent failure of the leading T cell based HIV vaccine in human efficacy trials. Even if such an immunogen can be developed, most animal model studies indicate that high serum neutralizing concentrations of bNAbs are required to provide significant benefit in typical protection experiments. One possible exception is provided by the anti-glycan bNAb 2G12, which has been reported to protect macaques against CXCR4-using SHIV challenge at relatively low serum neutralizing titers. Here, we investigated the ability of 2G12 administered intravenously (i.v.) to protect against vaginal challenge of rhesus macaques with the CCR5-using SHIVSF162P3. The results show that, at 2G12 serum neutralizing titers of the order of 1:1 (IC90), 3/5 antibody-treated animals were protected with sterilizing immunity, i.e. no detectable virus replication following challenge; one animal showed a delayed and lowered primary viremia and the other animal showed a course of infection similar to 4 control animals. This result contrasts strongly with the typically high titers observed for protection by other neutralizing antibodies, including the bNAb b12. We compared b12 and 2G12 for characteristics that might explain the differences in protective ability relative to neutralizing activity. We found no evidence to suggest that 2G12 transudation to the vaginal surface was significantly superior to b12. We also observed that the ability of 2G12 to inhibit virus replication in target cells through antibody-mediated effector cell activity in vitro was equivalent or inferior to b12. The results raise the possibility that some epitopes on HIV may be better vaccine targets than others and support targeting the glycan shield of the envelope.