TAG HIV Basic Science, Vaccines, and Cure Project Blog

Does HIV Unleash Endogenous Retroviral Elements?

Two interesting presentations on the first day of CROI related to human endogenous retroviruses (HERVs). HERVs are essentially fossil remnants of retroviruses that humans encountered many millennia ago; they can no longer replicate but rather have incorporated into human DNA and now make up a surprising 8% of our genome. Some rare HERV sequences (e.g. HERV-K) exist as full length proviruses but fragments known as human endogenous retrotransposable elements (HERE) are more common. Human cells have restriction factors like the recently discovered APOBEC proteins which suppress retroviral activity and these factors are believed to keep endogenous retroviral sequences dormant. However, it is known that HIV has evolved mechanisms (such as the viral protein Vif) which disable host restriction factors. This led researcher Brad Jones from the University of Toronto to wonder if perhaps HIV infection of CD4 cells would awaken endogenous retroviral sequences and allow them to become active.

When Jones looked at HIV-infected primary CD4 T cell lines in vitro, this is exactly what he found. These cells accumulated increasing numbers of genomic copies of the HEREs that Jones tested for (AluSX, LINE-1 and segments of HERV-K), but uninfected CD4 cells did not. To take a preliminary look at whether such events occur in vivo, Jones searched databases of HIV sequences and found a primary HIV isolate into which a HERE (LINE-1) had become inserted, strongly suggesting that HIV and LINE-1 can be active in the same cell.

Further supporting Jones's reasoning, Keith Garrison from UCSF presented a poster (#457) showing that people with HIV have higher levels of HERV-K genetic material in their blood compared to uninfected controls. Furthermore, T cell responses to HERV-K protein fragments (peptides) could be detected in individuals with primary HIV infection but not in uninfected controls.  There was even a weak but statistically significant inverse correlation between HERV-K-specific T cell responses and HIV viral load (r²=0.11, p=0.03). 

These findings may suggest novel strategies for HIV vaccines and immunotherapies. HIV's notorious ability to mutate makes it a difficult, moving target for immune responses. But endogenous retroviral proteins cannot mutate, and if they are only expressed in HIV-infected cells it is possible that triggering immune responses against these proteins with a vaccine would provide another means of targeting and eliminating HIV-infected cells.

Brad Jones's presentation will soon be available online as part of the Monday webcast of the CROI session entitled "Host-Cell Regulation of Viral Replication" (Monday Feb 26, 10:00 AM). Abstracts and PDFs of poster presentations will start becoming available online on the conference website after the meeting ends on February 28.

Conference on Retroviruses & Opportunistic Infections

The 14th Conference on Retroviruses & Opportunistic Infections (CROI) kicked off yesterday in Los Angeles. Webcasts from the conference are quickly being placed online, many of yesterday's sessions are already available.

Acute Infection is a Major Contributor to HIV Transmission

A new study and associated commentary are available free online from the Journal of Infectious Diseases:

The Journal of Infectious Diseases 2007;195:000

Primary HIV Infection, Phylogenetics, and Antiretroviral Prevention

Deenan Pillay and Martin Fisher

The Journal of Infectious Diseases 2007;195:000

High Rates of Forward Transmission Events after Acute/Early HIV-1 Infection

Bluma G. Brenner,1 Michel Roger,2 Jean-Pierre Routy,3 Daniela Moisi,1 Michel Ntemgwa,1 Claudine Matte,2 Jean-Guy Baril,4 Réjean Thomas,5 Danielle Rouleau,2 Julie Bruneau,6 Roger Leblanc,7 Mario Legault,8 Cecile Tremblay,9 Hugues Charest,10 Mark A. Wainberg,1 and the Quebec Primary HIV Infection Study Groupa

1McGill AIDS Centre–Jewish General Hospital, 2Centre Hospitalier de Université de Montréal (CHUM)–Hôpital Notre-Dame, 3McGill University Health Centre, 4Clinique Médicale du Quartier Latin, 5Clinique Médicale l'Actuel, 6CHUM–Hôpital St. Luc, 7Clinique Médicale Goldberg, LeBlanc, & Rosengren, 8Fonds de la Recherche en Santé du Québec–SIDA Network, 9CHUM–Hôtel Dieu, and 10Institut National de Santé Publique du Québec, Montreal, Canada

Background: A population-based phylogenetic approach was used to characterize human immunodeficiency virus (HIV)–transmission dynamics in Quebec.

Methods: HIV-1 pol sequences included primary HIV infections (PHIs; <6 months after seroconversion) from the Quebec PHI cohort (1998–2005; n = 215) and the provincial genotyping program (2001–2005; n = 481). Phylogenetic analysis determined sequence interrelationships among unique PHIs (n = 593) and infections from untreated (n = 135) and treated (n = 660) chronically infected (CI) potential transmitter populations (2001–2005). Clinical features, risk factors, and drug resistance for clustered and nonclustered transmission events were ascertained.

Results: Viruses from 49.4% (293/593) of PHIs cosegregated into 75 transmission chains with 2–17 transmissions/cluster. Half of the clusters included 2.7 ± 0.8 (mean ± SD) transmissions, whereas the remainder had 8.8 ± 3.5 transmissions. Maximum periods for onward transmission in clusters were 15.2 ± 9.5 months. Coclustering of untreated and treated CIs with PHIs were infrequent (6.2% and 4.8%, respectively). The ages, viremia, and risk factors were similar for clustered and nonclustered transmission events. Low prevalence of drug resistance in PHI supported amplified transmissions at early stages.

Conclusions: Early infection accounts for approximately half of onward transmissions in this urban North American study. Therapy at early stages of disease may prevent onward HIV transmission.

Initial Events in Vaginal HIV Transmission

An open access article just published in the journal Immunity offers new insights into the vaginal transmisison of HIV. A group of researchers from the University of Washington report the development of a human organ culture system that allows the behavior of HIV to be tracked in intact, viable vaginal outer epithelium. The researchers, led by Florian Hladik, use the system to show that HIV initially targets two different cell types: CD4 T cells, which it enters by direct binding to CD4 and the CCR5 coreceptor leading to productive infection, and Langerhans cells, which it enters via a process called endocytosis (in which the cell draws in material from outside). After HIV is endocytosed by the Langerhans cells it remains viable for several days in an internal compartment of the cell called the cytoplasm, but productive infection does not ensue. Previous research had suggested that HIV infection of CD4 T cells occurs only after transfer of virus from Langerhans cells but this study demonstrates clearly that the virus can enter both cell types simultaneously. About half the CD4 T cells from five tissue donors showed direct binding of virus and this proportion was reduced to ~6% in the presence of antibodies to CCR5 and CD4.

Langerhans cells are antigen-presenting cells (APCs) and their job is to transport foreign material to the lymph nodes for presentation to T cells, so these data also support previous evidence suggesting that APCs can act as Trojan horses that bring HIV into contact with T cells during the process of antigen presentation. Hladik and colleagues found that both infected CD4 T cells and Langerhans cells carrying HIV migrated out of the vaginal epithelium. This finding strongly suggests that, in vivo, these cells are capable of migrating to lymphoid tissue where APC/CD4 T cell interactions and CD4 T cell activation amplify and spread HIV infection systemically.

The researchers conclude by noting that the results have implications for strategies aiming to block vaginal transmission of HIV. Such approaches will likely need to try and block uptake of virus by Langerhans cells in addition to preventing entry into CD4 T cells.

Elite Suppression - Immunological Control of HIV?

A subset of HIV-infected individuals do not show any signs of disease progression and maintain normal CD4 T cell counts over many years of follow up; they are typically referred to as long-term non-progressors (LTNPs). Therapy is usually unnecessary in such individuals because the immunologic indicators that are used to guide initiation of antiretroviral treatment (the CD4 T cell count and/or percentage) remain stable. The advent of viral load tests has revealed that a subset of LTNPs maintain incredibly strict suppression of HIV replication, rarely experiencing viral loads above 50 copies/mL (the limit of detection of the test). This subset of LTNPs have been dubbed “elite suppressors.” One potential explanation of this phenomenon is that such individuals are infected with weak of defective viruses; a number of studies have identified mutations in HIV that may be associated with LTNP status.

In a study published in the latest issue of the Journal of Virology, Bob Siliciano and colleagues explore this question in a small group of 10 elite suppressors. The researchers successfully cultured virus from four of the individuals, ending up with six HIV isolates that could be assessed in detail. No known mutations affecting viral fitness were seen and all six viral isolates replicated as well as laboratory strains in vitro. All the viruses – both the isolates and the lab strains - could also replicate in CD4 T cells taken from the elite suppressors. Due to their inability to identify viral factors that might be explain the elite suppressor status of these individuals, the authors conclude by stating: “our results suggest that long-term immunologic control of fully pathogenic HIV-1 is possible.”

Journal of Virology, March 2007, p. 2508-2518, Vol. 81, No. 5

Isolation and Characterization of Replication-Competent Human Immunodeficiency Virus Type 1 from a Subset of Elite Suppressors

Joel N. Blankson,1 Justin R. Bailey,1 Seema Thayil,1 Hung-Chih Yang,1 Kara Lassen,1 Jun Lai,1,2 Shiv K. Gandhi,1 Janet D. Siliciano,1 Thomas M. Williams,3 and Robert F. Siliciano1,2

Department of Medicine, Johns Hopkins University School of Medicine,1 Howard Hughes Medical Institute, Baltimore, Maryland 21205,2 Department of Pathology, University of New Mexico School of Medicine, and Tricore Reference Laboratories, Albuquerque, New Mexico 871313

Received 3 October 2006/ Accepted 28 November 2006

Elite suppressors (ES) are untreated human immunodeficiency virus type 1 (HIV-1)-infected individuals who control viremia to levels below the limit of detection of current assays. The mechanisms involved in this control have not been fully elucidated. Several studies have demonstrated that some ES are infected with defective viruses, but it remains unclear whether others are infected with replication-competent HIV-1. To answer this question, we used a sensitive coculture assay in an attempt to isolate replication-competent virus from a cohort of 10 ES. We successfully cultured six replication-competent isolates from 4 of the 10 ES. The frequency of latently infected cells in these patients was more than a log lower than that seen in patients on highly active antiretroviral therapy with undetectable viral loads. Full-length sequencing of all six isolates revealed no large deletions in any of the genes. A few mutations and small insertions and deletions were found in some isolates, but phenotypic analysis of the affected genes suggested that their function remained intact. Furthermore, all six isolates replicated as well as standard laboratory strains in vitro. The results suggest that some ES are infected with HIV-1 isolates that are fully replication competent and that long-term immunologic control of replication-competent HIV-1 is possible.

Valproic Acid Fails to Deplete HIV Reservoirs

Back in 2005, a study was published in The Lancet suggesting that an anti-seizure medication – valproic acid – might help deplete the reservoir of latently infected CD4 T cells in people with HIV. Although the data were derived from only three people, The Lancet got more than a little carried away, placing a quote from the paper on its cover stating this may be a step toward a cure. Much press attention ensued.

Now, in the online first section of the Journal of Infectious Diseases, comes news that valproic acid appears to exert no such effect. Janet Siliciano, a researcher with extensive experience studying latently infected CD4 T cells, evaluated their numbers in nine people with HIV with well-suppressed viral load who were receiving valproic acid for neurological or psychiatric conditions. Siliciano reports that “Latently infected cells were readily detectable in all patients studied and did not in general decrease over time in a given patient. The estimated decay of the latent reservoir was extremely slow and was similar to that previously reported for patients receiving HAART alone.”

Abstract:

The Journal of Infectious Diseases 2007;195:000

BRIEF REPORT

Stability of the Latent Reservoir for HIV-1 in Patients Receiving Valproic Acid

Janet D. Siliciano,1 Jun Lai,1 Marc Callender,1 Eleanor Pitt,1 Hao Zhang,2 Joseph B. Margolick,2 Joel E. Gallant,1 Joseph Cofrancesco, Jr.,1 Richard D. Moore,1 Stephen J. Gange,3 and Robert F. Siliciano1,4

1Department of Medicine, Johns Hopkins University School of Medicine, Departments of 2Molecular Microbiology and Immunology and 3Epidemiology, Johns Hopkins Bloomberg School of Public Health, and 4Howard Hughes Medical Institute, Baltimore, Maryland

(See the editorial commentary by Schooley and Mellors, on pages XXX–XX.)

In light of a recent report that short-term treatment with valproic acid (VA) might accelerate the decay of the latent reservoir for HIV-1 in patients receiving combination therapy and allow eventual eradication of the infection, we studied patients with prolonged suppression of viremia who were receiving combination therapy and who had also been receiving chronic VA therapy for neurological or psychiatric conditions. Latently infected cells were readily detected in all patients at levels comparable to those seen in patients receiving combination therapy alone. We conclude that the clinical use of VA has no ancillary effect on the decay of the latent reservoir.

More Gut Rumblings

The February 1 issue of the journal Blood contains two new studies and a commentary that relate to recent theories on gut CD4 depletion and HIV pathogenesis. The most prominent of these theories holds that CCR5-expressing CD4 T cells in the gut are eliminated by the direct cytopathic effects of HIV within weeks of infection, setting the stage for the subsequent development of AIDS. In one of the new papers, Ivona Pandrea and colleagues (including Andrew Lackner and Ron Veazey, two leading proponents of the theory that gut CD4 depletion is central to HIV pathogenesis) demonstrate that natural hosts of SIV infection (sooty mangabeys, African green monkeys, mandrills, sun-tailed monkeys and chimpanzees) have a smaller proportion of CD4 T cells expressing CCR5 compared to primate species that suffer pathogenic consequences when infected with HIV or SIV (humans, rhesus macaques, pigtail macaques and baboons). This finding held true in both peripheral blood and gut-associated lymphoid tissue. The authors argue that this is likely a result of prolonged co-evolution with SIV and accounts for the typical lack of pathogenicity of the virus in its natural hosts.

The accompanying commentary by Mario Roederer and Joseph Mattapallil also states (in reference to the study): "low CCR5 expression was associated with better protection and preservation of gut CD4 T cells." However, the Pandrea paper contains no such data; it only reports the percentage of CCR5-expressing CD4 T cells in the gut of SIV-infected sooty mangabeys and African green monkeys, not the percentage or absolute number of gut CD4 T cells. Furthermore, the same group of researchers presented data at last year's CROI meeting showing that both SIV-infected sooty mangabeys and African green monkeys experience an apparent severe depletion of CD4 T cells from the gut despite the fact that they rarely develop signs of immunodeficiency and - as outlined in the new study - have few CCR5-expressing CD4 T cells in the gut.

In the second paper, it is reported that neonatal SIV-infected macaques experience a depletion of memory CD4 T cells from the gut similar to that seen in adult macaques. The authors note that finding high levels of memory CD4 T cells (as defined by expression of the molecules CD95 and CD28) in the gut was unexpected and speculate that they develop in response to environmental antigens in utero, as they are present even at birth. Another possibility might be that the definition of memory CD4 T cells obtained from studies of peripheral blood cannot straightforwardly be applied to CD4 T cells from the gut. Immunologists typically divide memory CD4 T cells into effector memory or central memory subsets, but the authors of this paper describe the cells being lost from the gut in these animals as "'activated' central memory." The reason this unusual term is employed is that CD4 T cells in the gut are typically highly activated and have a number of markers and properties that distinguish them from CD4 T cells sampled from other sites. As yet, not even specialists in gut immunology fully understand the reason for these distinguishing features, which perhaps suggests that proponents of gut CD4 depletion theories need to be circumspect in their attempts to categorize the type of CD4 T cell that is being lost. Two recent reviews - one by Zvi Grossman and colleagues, the other by John Zaunders and Anthony Kelleher - have argued that the definition of gut CD4 T cells in these studies as memory cells is misleading, since they are unlikely to be representative of the long-lived central memory CD4 T cell pool. Highly activated CD4 T cells, they point out, are mostly destined to undergo activation-induced cell death whether they are infected or not.

The outstanding questions regarding the role of gut CD4 T cells in HIV pathogenesis will need to be answered by future research. What is somewhat disheartening about this trio of papers is the extent to which the author's interpretation of the data is conveyed as fact. This is a road that prior purveyors of pathogenesis theories have gone down (such as David Ho with his "tap and drain" model of CD4 T cell depletion), and it has not served the field of HIV research well. Until gut immunology in both health and disease is better understood, a degree of caution regarding the interpretation of these and similar papers may be appropriate.

Blood, 1 February 2007, Vol. 109, No. 3, pp. 854

CCR5 and HIV: the less, the better

Mario Roederer, and Joseph Mattapallil

Two reports in this issue of Blood underscore the central importance of the regulation of chemokine receptor CCR5 expression to the pathogenic infection of SIV and associated destruction of gut mucosa in nonnatural hosts. CCR5 is the obligate coreceptor for infection by SIV and most transmitted forms of HIV; humans who lack CCR5 expression (CCR5{Delta}32) are highly resistant to HIV infection. Now it appears that a selection for low expression of CCR5 in natural hosts of SIV has evolved as one mechanism to escape the pathogenic consequences of infection.

Blood, 1 February 2007, Vol. 109, No. 3, pp. 1069-1076.
DOI 10.1182/blood-2006-05-024364.

Paucity of CD4+CCR5+ T cells is a typical feature of natural SIV hosts

Ivona Pandrea1, Cristian Apetrei1, Shari Gordon2,3, Joseph Barbercheck1, Jason Dufour1, Rudolf Bohm1, Beth Sumpter3, Pierre Roques4, Preston A. Marx1, Vanessa M. Hirsch5, Amitinder Kaur6, Andrew A. Lackner1, Ronald S. Veazey1, and Guido Silvestri2,3

1 Tulane National Primate Research Center, Covington, LA; 2 Department of Pathology, University of Pennsylvania, Philadelphia, PA; 3 Yerkes National Primate Research Center, Atlanta, GA; 4 Centre International de Recherches Medicales, Franceville, Gabon; 5 Laboratory of Molecular Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD; 6 New England Primate Research Center, Southborough, MA

In contrast to lentiviral infections of humans and macaques, simian immunodeficiency virus (SIV) infection of natural hosts is nonpathogenic despite high levels of viral replication. However, the mechanisms underlying this absence of disease are unknown. Here we report that natural hosts for SIV infection express remarkably low levels of CCR5 on CD4+ T cells isolated from blood, lymph nodes, and mucosal tissues. Given that this immunologic feature is found in 5 different species of natural SIV hosts (sooty mangabeys, African green monkeys, mandrills, sun-tailed monkeys, and chimpanzees) but is absent in 5 nonnatural/recent hosts (humans, rhesus, pigtail, cynomolgus macaques, and baboons), it may represent a key feature of the coevolution between the virus and its natural hosts that led to a nonpathogenic infection. Beneficial effects of low CCR5 expression on CD4+ T cells may include the reduction of target cells for viral replication and a decreased homing of activated CD4+ T cells to inflamed tissue. 

Blood, 1 February 2007, Vol. 109, No. 3, pp. 1174-1181.
DOI 10.1182/blood-2006-04-015172.

Massive infection and loss of CD4+ T cells occurs in the intestinal tract of neonatal rhesus macaques in acute SIV infection

Xiaolei Wang1, Terri Rasmussen1, Bapi Pahar1, Bhawna Poonia1, Xavier Alvarez1, Andrew A. Lackner1, and Ronald S. Veazey1

1 Tulane National Primate Research Center, Tulane University School of Medicine, Covington, LA

Rapid, profound, and selective depletion of memory CD4+ T cells has now been confirmed to occur in simian immunodeficiency virus (SIV)–infected adult macaques and human immunodeficiency virus (HIV)–infected humans. Within days of infection, marked depletion of memory CD4+ T cells occurs primarily in mucosal tissues, the major reservoir for memory CD4+ T cells in adults. However, HIV infection in neonates often results in higher viral loads and rapid disease progression, despite the paucity of memory CD4+ T cells in the peripheral blood. Here, we examined the immunophenotype of CD4+ T cells in normal and SIV-infected neonatal macaques to determine the distribution of naive and memory T-cell subsets in tissues. We demonstrate that, similar to adults, neonates have abundant memory CD4+ T cells in the intestinal tract and spleen and that these are selectively infected and depleted in primary SIV infection. Within 12 days of SIV infection, activated (CD69+), central memory (CD95+CD28+) CD4+ T cells are marked and persistently depleted in the intestine and other tissues of neonates compared with controls. The results in dicate that "activated" central memory CD4+ T cells are the major target for early SIV infection and CD4+ T cell depletion in neonatal macaques.   

PD-1 Papers Proliferate

In the advance online section of the journal Blood, a group of researchers based in Beijing, China present results confirming that expression of the molecule PD-1 is a useful marker for HIV-specific CD8 T cell exhaustion. The investigators compared expression of the molecule on  HIV-specific CD8 T cells from people with progressing infection versus long-term non-progressors and found significant differences between the two groups. Similar to previously reported results, the authors also found that blocking PD-1/PD-L1 interactions in vitro appeared to restore potentially important CD8 T cell functions, such as the ability to proliferate in response to HIV. While the authors note that these findings may have therapeutic implications, there is increasing concern among researchers that non-specific attempts to block PD-1/PD-L1 interactions in vivo could provoke catastrophic autoimmunity. For example, unpublished research from Rafi Ahmed's group has shown that blocking PD-L1 during acute LCMV infection leads to fatal immunopathology in mice. Currently, several groups of researchers are planning to carefully evaluate the effects of anti-PD-1 strategies in animal models, including SIV-infected macaques and HCV-infected chimpanzees. Murine researchers are also investigating the effects of blocking interactions between PD-1 and it's second ligand, PD-L2, which is expressed on a far more limited range of cells than PD-L1 (primarily dendritic cells and macrophages), in the hopes that this approach may offer a safer means to enhance the function of exhausted T cells.

Blood First Edition Paper, prepublished online February 1, 2007; DOI 10.1182/blood-2006-09-044826.

PD-1 upregulation is correlated with HIV-specific memory CD8+ T-cell exhaustion in typical progressors, but not in long-term non-progressors

Ji-Yuan Zhang, Zheng Zhang, Xicheng Wang, Jun-Liang Fu, Jinxia Yao, Yanmei Jiao, Liangen Chen, Hui Zhang, Jianan Wei, Lei Jin, Ming Shi, George Fu Gao, Hao Wu, and Fu-Sheng Wang*

Institute of Microbiology, Chinese Academy of Sciences (CAS), Beijing, China
Research Center of Biological Therapy, Beijing, China
Department of Infectious Diseases, Beijing You-An Hospital Affiliated to Capital University of Medical Science, Beijing, China
HIV/AIDS Research Center, Beijing Guang-An-Men Hospital, Beijing, China

The immunoreceptor PD-1 is significantly upregulated on exhausted CD8+ T cells during chronic viral infections like HIV-1. However, it remains unknown whether PD-1 expression on CD8+ T cells differs between typical progressors (TP) and long-term non-progressors (LTNP). In this report, we examined PD-1 expression on HIV-specific CD8+ T cells from 63 adults with chronic HIV infection. We found that LTNP exhibited functional HIV-specific memory CD8+ T cells with markedly lower PD-1 expression. TP, in contrast, showed significantly up-regulated PD-1 expression that was closely correlated with a reduction in CD4 T-cell number and an elevation in plasma viral load. Importantly, PD-1 upregulation was also associated with reduced perforin and IFN-{gamma} production, as well as decreased HIV-specific effector memory CD8+ T-cell proliferation in TP but not LTNP individuals. Blocking PD-1/PD-L1 interactions efficiently restored HIV-specific CD8+ T-cell effector function and proliferation. Taken together, these findings confirm the hypothesis that high PD-1 upregulation mediates HIV-specific CD8+ T-cell exhaustion. Blocking PD-1/PD-L1 pathway may represent a new therapeutic option for this disease, and provide more insight into immune pathogenesis in LTNP individuals.