Current HIV Research (v.8, #8)

A recent phase III vaccine trial in Thailand has shown for the first time that a vaccine can prevent HIV-1 infection in humans [1]. Although the efficacy was modest, and the protection might have been short-lived, the results of this trial have raised hopes in the field of HIV-1 vaccine research after a number of failures. The vaccination protocol consisted in priming with a canarypox virus vector component expressing Gag, protease, and gp41 derived from an HIV-1 subtype B isolate and gp120 from a CRF01_AE virus, and boosting with envelope glycoproteins of isolates of subtype B and CRF01_AE. The genetic composition of the immunogens was guided by phylogenetic and molecular epidemiological studies of the viruses circulating in the targeted population, among which a CRF01_AE variant, introduced in Thailand the late 1980s, is the predominant clade, with a subtype B strain introduced around the same time, circulating as a minor genetic form, and by cross-neutralization studies indicating that the Thai CRF01_AE variant correlates with a neutralization serotype [2]. Whether the genetic relatedness between the vaccine immunogens and the locally circulating HIV-1 variants favored vaccine-mediated protection is the subject of ongoing studies. Whatever their results, the design of the immunogens of this vaccine and of others planned for future testing, in which the immunogens match the HIV-1 variants circulating among the targeted population, exemplify the growing and widely accepted idea that HIV-1 genetic diversity is an essential element to be taken into consideration in HIV-1 vaccine design, which is the central topic of the articles of this special issue of Current HIV Research. This idea has not always had a general support, with most earlier work failing to find correlations between antibody-mediated neutralization and HIV-1 subtypes, and most studies on T cell-mediated responses emphasizing the detection of cross-clade reactivities. Based on those studies, it was suggested that a HIV-1 vaccine design approach targeted to subtypes seemed and#x201C;unnecessary and without scientific foundationand#x201D; [3]. However, multiple subsequent studies have supported that there may be a correlation between HIV-1 clades and antibody-mediated neutralization (reviewed by van Gils and Schuitemaker) and have detected cell-mediated responses with preferential or specific intraclade reactivities (reviewed by McKinnon et al., who also underscore the importance of inducing cross-reactive CD8+ T cell responses for vaccine protection, and of using multiple immunological assays for their detection). The relevance of viral genetic diversity for vaccine efficacy is also supported by results obtained with other lentiviruses. A study with an attenuated equine infectious anemia virus (EIAV) vaccine in ponies found an inverse linear correlation between vaccine efficacy and increasing genetic divergence of the challenge virus gp90 (the EIAV envelope surface glycoprotein), with an amino acid distance of as little as 6and#x25; from the vaccine surface glycoprotein resulting in a 25and#x25; reduction in protection from disease, which increased to 50and#x25; when the divergence was 13and#x25; [4]. On the other hand, single-subtype feline immunodeficiency virus vaccines have been protective only against homologous or homologous-subtype in vitro-derived challenges [5]. Further knowledge on the significance of HIV-1 variants for immune responses relevant for vaccine protection in humans might come, apart from the vaccine trials themselves, from large population-based studies on the incidence of superinfection (reviewed by Chohan et al.) in areas where multiple clades circulate. The difficulty in detecting strong correlations between HIV-1 subtypes and immune responses may derive from multiple causes: a) large intrasubtype genetic distances, which may currently be similar to or greater than intersubtype divergence at the time of the origin of subtypes, with epitopes originally shared within clades being progressively lost through accumulation of mutations along subtype diversification; b) frequent intersubtype recombination in areas where multiple subtypes circulate, which may confound the results of cross-neutralization assays when envelope sequences of the isolates used in the assays or of those inducing the antibody responses are not fully characterized, as occurred in some of the earlier studies; c) cryptic dual infections with viruses of different clades, in which one of the variants remains undetected due to its low abundance, with immune responses induced by the minority variant being erroneously interpreted as cross-clade responses; d) the use of assays to detect cytotoxic T lymphocyte responses that may not reflect antiviral activity [6]. Therefore, it is important that these points be taken into consideration when conducting studies on HIV-1 cross-clade immune responses. Given the great degree of genetic divergence within HIV-1 subtypes, it may also be relevant to consider the intrasubtype phylogenetic structure in vaccine-related studies. Such structure is apparent in some geographic areas, where a large proportion of viruses branch within well supported intrasubtype clusters [7-9]. In other areas, extensive intrasubtype recombination may have blurred the distinction between clusters, which may not be apparent when employing the usual methods of phylogenetic analysis. In this regard, the detection of regional clustering of neutralization, as reported for subtype C isolates from South Africa [10], may reflect the existence of an underlying intrasubtype phylogenetic clustering (Thomson MM, unpublished results). The lower genetic diversity within intrasubtype clusters or variants, compared to subtypes, would be expected to be reflected in stronger correlations with immune responses (as observed with the Thai CRF01_AE variant in neutralization assays [2]), which may be of relevance for the genetic composition of the vaccine immunogens used in areas where such clusters or variants are circulating. Different strategies for immunogen design have been devised to broaden the immune responses effective across the wide diversity of HIV-1 variants. One of them is the use of polyvalent formulations consisting of cocktails of immunogens derived from different HIV-1 clades, similarly to other widely used vaccines of proven efficacy against other pathogens (reviewed by Lu et al.). Another is the use of immunogens with centralized (either ancestral, consensus, or center of tree) sequences, designed to minimize the genetic distances with a majority of circulating viruses within a clade (reviewed by Arenas and Posada, who underscore the importance of taking into account recombination and complex models of evolution for the design of such immunogens). Additional strategies include the use of computationally designed mosaic proteins assembled from natural sequences optimized to maximize coverage of the most common potential T-cell epitopes across HIV-1 clades [11], and of immunogens incorporating the most highly conserved regions of the HIV-1 proteome, in which escape mutations would be predicted to severely compromise virus viability [12-14]. These strategies are not mutually exclusive and could be used in combination. In rare HIV-1-infected individuals antibodies are produced possessing broad and potent neutralizing activity across HIV-1 clades (reviewed by Gonzalez et al.). Rational design of Env immunogens based on the epitopes targeted by such antibodies would be another strategy for broadening humoral responses elicited by HIV-1 vaccines. However, it is uncertain whether such a vaccine could induce these rare antibodies in the general population, if their elicitation depends on the particular primary B-cell repertoires present in a few individuals and on a lengthy affinity maturation process of the antibody response in the context of a chronic HIV-1 infection [15]. Given the lack of adequate animal models, definitive knowledge on the significance of HIV-1 genetic diversity for vaccine efficacy in humans will only come from large-scale vaccine trials in populations among which different HIV-1 variants (including subtypes, circulating recombinant forms, and intraclade variants) are circulating. However, cumulating evidence, as reviewed in this issue, supports the relevance of the correlations of HIV-1 genetic forms to immune responses for vaccine design. This constitutes a rapidly expanding area of research which will most likely be of key importance for the development of effective preventive vaccines against HIV-1.

The correlates of protection against HIV-1 infection or disease progression are still unknown which causes an immense challenge for HIV-1 vaccine design. Existing effective vaccines against other viruses generate antibodies that either block the initial infection or contribute to the eradication of the virus before it can cause disease. For HIV-1, a protective vaccine capable of eliciting protective neutralizing antibodies does not exist and the difficulties for the generation of such a vaccine are multiple. Conserved elements on the viral envelope glycoprotein, the target of HIV specific neutralizing antibodies, seem to be poorly immunogenic and attempts to generate an immunogen that can elicit broadly reactive neutralizing antibodies have remained largely without success. In addition, the envelope of HIV-1 is highly variable with respect to amino acid sequence, length of the variable loops, and glycosylation pattern. To cope with the high sequence variation, vaccine-elicited clade-specific neutralizing antibodies have been suggested as an attractive alternative and recent studies have revealed some evidence for the existence of HIV-1 clade-specific humoral immune responses. Here, we will review these recent findings and hypothesize on the nature of clade-specific humoral immunity also in light of their relevance for HIV-1 vaccine development.

HIV-Specific T Cells: Strategies for Fighting a Moving Target by Lyle R. McKinnon, Rupert Kaul, Melissa Herman, Francis A. Plummer, T. Blake Ball (587-595).
HIV vaccine development faces many hurdles, including the failure of empirical approaches, an incomplete understanding of protective immunity, and the extreme genetic diversity of HIV-1. HIV is a moving target in at least two key ways: 1) within an infected individual, years of evolution lead to the formation of quasispecies, and selection of variants with increased fitness, and 2) during the course of the pandemic, subtypes change in frequency as they are transmitted from host to host. In spite of this, CD8+ T cells are often able to overcome HIV diversity, leading to relatively high levels of cross-reactive and cross-clade responses. Recent research suggests that the cross-reactivity of HIV-specific CD8+ T cell responses should be evaluated comprehensively, using multiple immunological assays (including those that correlate best with protective immunity), and taking into account subtle differences in epitopic variation, presenting HLA allele, and cognate TCR that all influence recognition and escape. In addition, although escape and cross-reactivity are often predictable, important differences can be present, particularly in the setting of multiple and different clades. Finally, strategies to optimize the induction of protective, cross-reactive T cells, and towards the likely infecting strain in the mucosa where exposure occurs and opportunities to prevent infection are greatest, are urgently needed. Though some cues can be found from observational studies, more in depth analyses of past and ongoing HIV vaccine trials will be needed to know if and how HIV genetic diversity can be overcome by vaccine-induced T cells.

HIV-1 Superinfection and its Implications for Vaccine Design by Bhavna H. Chohan, Anne Piantadosi, Julie Overbaugh (596-601).
HIV-1 superinfection, which refers to a subsequent HIV-1 infection from a different source partner after the first HIV-1 infection is established, has now been well documented in multiple populations. Some studies suggest that the risk of superinfection may be close to that of initial infection, suggesting that the immunity induced by chronic HIV-1 infection may not be adequate to confer protection from another HIV-1 strain. Detailed studies that examined immune responses in individuals who became superinfected generally support this hypothesis, but such studies have been limited. Indeed, superinfection represents one of the few settings, apart from vaccine trials, where there is an opportunity to gain insights into the role of HIV-specific immunity in protection in humans, and this should be exploited. Likewise, studies of superinfection in HIV-1 positive individuals on antiretroviral therapy who continue to be exposed to HIV could provide insight into the role of antiretroviral treatment in protecting from HIV-1 infection, a concept that is also being explored for its potential to prevent a first HIV-1 infection. To address these questions, larger population-based studies that define the incidence and timing of superinfection and include collection of samples for immunological studies are needed.

Broadly Neutralizing Antibodies and their Significance for HIV-1 Vaccines by Nuria Gonzalez, Amparo Alvarez, Jose Alcami (602-612).
Despite extensive efforts, a preventive HIV vaccine has not yet been obtained and remains the main challenge in the field of AIDS research. Empirical approaches which have proved successful for many vaccines are not sufficient to tackle HIV-1 and new strategies to design effective preventive AIDS vaccines are critical. To this aim, further understanding of the mechanisms of action of neutralizing antibodies is required. In this review we summarize our current knowledge on the structure of the gp160 viral envelope and the dynamics of viral entry, the evolution of humoral response in HIV-infected patients and the mechanisms of viral escape. Finally, we describe the few neutralizing antibodies with activity against a broad spectrum of circulating HIV strains and their relevance in the design of new candidates to HIV-1 vaccines.

Computational Design of Centralized HIV-1 Genes by Miguel Arenas, David Posada (613-621).
The extreme genetic diversity of the HIV-1 remains as a daunting challenge for the development of an effective AIDS vaccine. One strategy for creating a single vaccine that protects against the HIV-1 expanding population is to reconstruct centralized immunogenic sequences that minimize the genetic distance to circulating strains that the vaccine is targeting. Such centralized genes can be estimated with inferred consensus, ancestral and center-of-tree sequences. Although the increased breadth of antibody and T-cell responses induced by the centralized vaccines to date are encouraging, they are modest and may only be partly effective in combating HIV-1. One of the reasons of this limited success might be that several features of HIV-1 molecular evolution have not been yet taken into account in the design of these centralized vaccines, the most important likely being its high recombination rate and complex nucleotide substitution process. Here we describe evolutionary methodologies for the inference of centralized HIV-1 genes, with particular focus on the sources of error introduced by recombination and the model of evolution, in order to foster the development of more effective immunogens before synthesis and assessment in the lab, and final testing in AIDS vaccine trials.

Polyvalent AIDS Vaccines by Shan Lu, Jill M. Grimes Serrano, Shixia Wang (622-629).
A major hurdle in the development of a global HIV-1 vaccine is viral diversity. For close to three decades, HIV vaccine development has focused on either the induction of T cell immune responses or antibody responses, and only rarely on both components. After the failure of the STEP trial, the scientific community concluded that a T cell-based vaccine would likely not be protective if the T cell immune responses were elicited against only a few dominant epitopes. Similarly, for vaccines focusing on antibody responses, one of the main criticisms after VaxGen's failed Phase III trials was on the limited antigen breadth included in the two formulations used. The successes of polyvalent vaccine approaches against other antigenically variable pathogens encourage implementation of the same approach for the design of HIV-1 vaccines. A review of the existing HIV-1 vaccination approaches based on the polyvalent principle is included here to provide a historical perspective for the current effort of developing a polyvalent HIV-1 vaccine. Results summarized in this review provide a clear indication that the polyvalent approach is a viable one for the future development of an effective HIV vaccine.

Barriers and Facilitators for Adherence to Antiretroviral Therapy in Papua New Guinea by A. Kelly, H. Worth, N. Man, S. Nosi, R. Emori, A. Mek, F. Akuani, M. Kupul, B. Kepa, L. Walizopa, L. Pirpir, B. Cangah, P. Siba, A. Frankland, P. Rawstorne (630-637).
Papua New Guinea (PNG) is in a phase of scaling up access to antiretroviral therapy (ART), and adherence to the newly available drug therapy is becoming an important issue. This paper examines adherence to ART in a sample of 374 HIV-positive people in six provinces in PNG. Participants were recruited to the study using non-probability sampling. Sixty-two and#x25; of participants reported complete adherence (no missed or late doses in the past week) and 79and#x25; reported not missing any doses in the last week. Revival church members were significantly more likely to report having missing a treatment dose(s) (66and#x25;). Those living in the Highlands and those attending Catholic health clinics were significantly more likely to be adherent to their treatment. Age, gender, marital status, education level and employment type did not show significant association with treatment adherence. Adherence rates in PNG are not alarming, indicating that people with HIV can adhere to treatment despite the challenges of living in PNG.

HIV-1 Nef Protein Visits B-Cells via Macrophage Nanotubes: A Mechanism for AIDS-Related Lymphoma Pathogenesis? by Susanna L. Lamers, Gary B. Fogel, Leanne C. Huysentruyt, Michael S. McGrath (638-640).
This letter refers to the recent demonstration that HIV-1 infected macrophages form specialized conduits that connect to B-cells (1). The conduit selectively transports the HIV-1 nef protein, providing nef with numerous means to interfere with cellular processes. Currently, no consideration of the connection between the conduit and the development of AIDS-related lymphoma (ARL) has been offered. ARL is one of the primary causes of death in the HIV-infected population and is related to B-cell proliferation and activation. In this letter we discuss several studies that link HIVinfected macrophages and specific forms of the nef protein to the development of ARL. The conduits discovered by Xu et al. [1] may lead to a better understanding of how HIV infection results in lymphomagenesis.

Use of Highly Active Antiretroviral Therapy is Increasing in HIV Positive Severe Drug Users by G. Vallecillo, A. Sanvisens, E. Martinez, M. Torrens, F. Bolao, J. Tor, R. Muga (641-648).
Background: Substance abuse greatly impacts the effectiveness of highly active antiretroviral therapy (HAART). We analyzed antiretroviral use in drug users positive for human immunodeficiency virus (HIV) that sought substance abuse treatment. Methods: This cross-sectional study recruited 705 patients HIV positive (74.6and#x25; men) between 1997 and 2007. Patients were grouped by calendar periods when different HAART regimens were available in Spain (p1: 1997-1999, n=299; p2: 2000-2003, n=249; and p3: 2004-2007, n=157). Results: The mean age at admission was 34 years; 94.7and#x25; had a past history of injection drug use (IDU) and 67.7and#x25; were current IDUs. The average CD4 cell count was 399 cells/and#956;L [interquartile range: 203-632 cells/and#956;L]. Lifetime prevalence of antiretroviral use was 59.4and#x25; (416/705; p1: 48.1and#x25;; p2: 64.6and#x25;; p3: 72.6and#x25;; p andlt; 0.05). The overall prevalence of antiretroviral use at admission was 40.7and#x25; (p1: 31.4and#x25;; p2: 41.0and#x25;; p3: 58.0and#x25;; p andlt; 0.05). In multivariate logistic regression analysis, age, calendar period, and non-IDU were predictors of antiretroviral use at admission. Among those taking antiretrovirals, 21.6and#x25; were on suboptimal HAART, mostly in the p1 group. Overall, 44.6and#x25; of patients were taking protease inhibitor and non-nucleoside reverse transcriptase inhibitor (PI-NNRTI), 21.9and#x25; were taking NRTI-NNRTI, and 9.4and#x25; were taking three NRTIs. Although not significant, the three-NRTI regimen was associated with CD4 > 350 cells/and#956;L and HIV RNA andlt; 400 copies/mL. Conclusions: HAART use is steadily increasing in HIV positive heavy drug users. However, part of this population remains antiretroviral therapy-naive despite advanced immunodeficiency. Interventions that focus on integrating substance abuse with HIV/AIDS treatments are needed.

Objectives: To compare rates of initial virological suppression and subsequent virological failure by Aboriginal ethnicity after starting highly active antiretroviral therapy (HAART). Methods: We conducted a retrospective cohort study of antiretroviral-naive HIV-patients starting HAART in January 1999-June 2005 (baseline), followed until December 31, 2005 in Alberta, Canada. We compared the odds of achieving initial virological suppression (viral load andlt; 500 copies/mL) by Aboriginal ethnicity using logistic regression and, among those achieving suppression, rates of virological failure (the first of two consecutive viral loads > 1000 copies/mL) by Aboriginal ethnicity using cumulative incidence curves and Cox proportional hazards models. Sex, injection drug use as an HIV exposure category (IDU), baseline age, CD4 cell count, viral load, calendar year, and HAART regimen were considered as potential confounders. Results: Of 461 study patients, 37and#x25; were Aboriginal and 48and#x25; were IDUs; 71and#x25; achieved initial virological suppression and were followed for 730.4 person-years. After adjusting for confounding variables, compared to non-Aboriginals with other exposures, the odds of achieving initial virological suppression were lower for Aboriginal IDUs (odds ratio (OR)=0.33, 95and#x25; CI=0.19-0.60, p=0.0002), non-Aboriginal IDUs (OR=0.30, 95and#x25; CI=0.15-0.60, p=0.0006), and Aboriginals with other exposures (OR=0.38, 95and#x25; CI=0.21-0.67, p=0.0009). Among those achieving suppression, Aboriginals experienced higher virological failure rates and#x2265;1 year after suppression (hazard ratio=3.35, 95and#x25; CI=1.68-6.65, p=0.0006). Conclusions: Future research should investigate adherence among Aboriginals and IDUs treated with HAART and explore their treatment experiences to assess ways to improve outcomes.