Current HIV Research - Volume 5, Issue 6, 2007

Each year, more than 5 million individuals are newly infected with HIV-1 [1]. To address this, the development of an HIV/AIDS vaccine has remained one of the highest priorities in biomedical research for the last 25 years. During this time, we have witnessed some success with the development of antiretroviral therapies capable of lowering the viral loads and decreasing the severity of disease. While treatment has extended the time course of infection, it has not resulted in a cure and/or prevention of HIV-1 infection. On the other hand, a large number of vaccine candidates have been tested with less than stellar success. Over the last two decades, researchers have focused almost exclusively on vaccines that exploit either cellular or humoral immunity as a goal for AIDS vaccine development. After more than two decades of limited success with vaccines, most researchers now believe that the goal of an AIDS vaccine should be to elicit as robust and broadly active cellular and humoral immunity as possible, thereby maximizing the potential for protection from variant HIV-1 strains by different routes of exposure. Researchers are returning focus back to the importance of antibody-mediated neutralization as a prime determinant in the protection against HIV-1 infection. One of the challenges for the development of an HIV/AIDS vaccine remains the great diversity of sequences present in viral isolates, particularly those in Env. HIV-1 isolates are divided into three different groups based on sequence diversity: Group M, N and O [2]. Isolates of group M (main) comprise the majority of worldwide infections and these viruses are further subdivided into nine different clades A-D, F-H, J, and K. These subtypes are separated by approximately 25-35% amino acid divergence in the surface glycoprotein Env. In addition, there are also circulating recombinant forms (CRFs) which are mosaics of two or more different viral isolates found in at least three epidemiologically independent individuals [2]. Therefore, the elicitation of an antibody response with broad neutralizing activity against primary isolates of multiple HIV-1 subtypes continues to be a desired characteristic of candidate HIV-1 vaccines. Standardized criteria for measuring the immune responses are critical and are being established. The use of new methodologies and testing requirements will provide a stronger environment for the development of broadly protective HIV-1 vaccines. In order to develop a successful AIDS vaccine depends on strategies to overcome the ability of the virus to evade humoral immunity at both the mucosal and systemic sites. This issue of Current HIV Research focuses on several aspects of our present scientific understanding of HIV neutralization. The structure and epitopes of the envelope glycoprotein are reviewed in association with design vaccine immunogens. Hu and Statamatos [3] provide an overview on Env modifications and their limitations vs. successes in vaccine design, while and Lin and Nara [4] discuss the design of core, functional Env modifications using immune dampening and immune refocusing strategies designed to counter immunodominant, decoy responses generated by the virus. Zwick and Burton provide an elegant review of the stoichiometry, kinetic, and thermodynamic parameters involved in neutralization [12]. Pinter describes the roles of Env variable regions important in HIV-1 neutralization that should be considered in vaccine development [5] and Burke and Barnett [13] provide a comparative overview of the described neutralizing epitopes in Env. The HIV-1 envelope is able to assume a variety of structures during binding to various cell receptors during the fusion and entry process, thereby exposing new epitopes to the immune system. DeVico describes the accessibility, antigenicity and immunogenicity of CD4-induced epitopes as possible vaccine immuongens [7]. Two reviews tackle the important issues regarding the diversity of the HIV-1 envelope. McKnight and Aasa-Chapman [6] describe the advantages of clade-specific versus clade-generic vaccines while Gao et al. [8] provide an overview of the use of centralized Env sequences to develop immunogens to elicit broadly reactive antibodies that neutralize isolates from all clades. Two reviews address the critical issue of HIV-1 transmissions across mucosal barriers....

So far, all efforts to engineer immunogens that would elicit broadly reactive anti-HIV neutralizing antibody responses have not been successful. In the past few years, however, key information on the structure of the epitopes recognized by several broadly reactive anti-HIV neutralizing antibodies (NAbs), the structures of these NAbs themselves and the molecular interaction between these NAbs and their epitopes has emerged, that promises to guide the design of better immunogens. In order to enhance the immunogenicity of conserved neutralization epitopes on Env, certain modifications such as variable loop-deletion, elimination of glycosylation sites, or epitope-repositioning, are being investigated. So far, however, all available data from immunization studies indicate that the effect such structural modifications have on Env immunogenicity is unpredictable. This implies that despite the significant progress made in elucidating the interaction of NAbs with their targets at the molecular level, a significant iterative effort is required to identify immunogens that would elicit the much anticipated broad anti-HIV neutralizing antibody responses.

To date HIV-1 vaccines have not been able to elicit potent, long lasting, and broadly neutralizing antibodies to the virus. Our knowledge of HIV envelope glycoprotein (Env) structure/function and the existence of a handful of broadly neutralizing antibodies is guiding rational immunogen design. We review here the potential targets on the HIV Env (the glycan shield, the CD4 binding site, the coreceptor binding site, Env fusion intermediates, and the membrane proximal region) and their associated rational immunogen design strategies. Moreover, we discuss immune dampening and immune refocusing strategies designed to counter immunodominant, decoy responses generated by the virus. In this regard, an immunogen design strategy of “in vitro de-evolution” is presented, which begins to distill the HIV Env to its most critical, core functional domains. While we are beginning to have some understanding as to where we would like out immune system to go, we find that our immune repertoire may actually have limits that preclude successful completion of the task at hand. The repertoire limits appear to be a byproduct of autoantibody tolerance mechanisms and the complex structural requirements for effective, potent broadly neutralizing antibodies. Nevertheless, the hope is that through novel insights and creative solutions that we will be able to design immunogens capable of eliciting broadly neutralizing antibodies to the HIV envelope glycoprotein.

A major focus of HIV-1 vaccine development has been directed towards a limited number of broadly conserved epitopes in the Envelope (Env) proteins that are sensitive neutralization targets in many primary isolates. However, evidence suggests that these epitopes are poorly immunogenic; similar antibodies are rarely produced by infected subjects, nor are they induced by various immunogens designed to express these epitopes. On the other hand, the major variable domains of Env are highly immunogenic; antibodies against these regions are common in sera of infected patients and easily generated upon immunization. Although these epitopes are extremely sensitive neutralization targets in some laboratory strains and primary isolates, the neutralization range of antibodies against these sites is limited. This review describes potent neutralization epitopes located in the variable regions of Env and discusses the bases for the limited neutralization breadth of antibodies against these targets. Strategies are discussed for using available information to design immunogens capable of exploiting the potential of these regions as vaccine targets.

The enormous diversity of the human immunodeficiency virus (HIV) has led to the idea that designing vaccines to specific geographic regions, or clades, could simplify the complexity of the task. Yet, despite the sequence diversity, all HIV viruses known to date interact with the same cellular receptors (CD4 and/or a coreceptor, CCR5 or CXCR4). In this review we examine the existing evidence to support a clade-specific vaccine strategy for induction of neutralising antibodies. We concentrate on lessons learnt from natural infection of humans. In short, the vast majority of studies to date indicate that neutralisation of HIV-1 is not clade specific. Potent sera tend to neutralise a range of heterologous viruses with no apparent clade preference, and none of the human neutralising monoclonal antibodies so far generated demonstrate significant clade preference. All but one of the most broadly neutralising antibodies are to functional regions involved in receptor interactions and plasma membrane fusion. Given these facts, we suggest that vaccine approaches that focus on ‘clade-specific’ and ‘clade-generic’ vaccines will logically converge on the same functionally conserved envelope structures. It still remains to be determined whether or not the task of designing a ‘clade-generic’ vaccine could be simplified by focusing on the viral envelopes with ‘transmitting phenotypes’.

The HIV surface glycoprotein, gp120, contains conserved and functional domains that exist within a viral envelope spike that is otherwise highly variable with respect to conformation, sequence and structure. Termed CD4-induced epitopes, these domains are stabilized on transition state gp120 structures as a consequence of CD4 receptor engagement. These nuggets of conservation naturally attract the attention of those seeking to develop vaccine and therapeutic strategies to fight infection by diverse HIV strains. However, an appreciation for the immunological relevance and practical value of CD4-induced epitopes is still evolving. This review covers the current findings related to the accessibility, antigenicity and immunogenicity of CD4-induced epitopes on gp120.

Centralized HIV-1 genes (consensus, most recent common ancestor and center of the tree) have recently been explored for induction of broadly reactive immune responses to overcome the extraordinary genetic diversity among HIV- 1 strains. Although all of these strategies are based on artificial sequences predicted by computer programs, they retain biological function, and use the CCR5 co-receptor for entry into target cells as transmitted HIV-1 Envs. Results from laboratory animals indicate that centralized immunogens are superior to many wild-type immunogens for inducing crosssubtype T and B cell immune responses. Structural modifications have improved the ability of consensus Envs to elicit antibody responses that neutralize a spectrum of HIV-1 Env pseudoviruses. However, the more difficult to neutralize tier 2 Env pseudoviruses are generally not neutralized well by anti-consensus Env antibodies, indicating the need for further modifications, new formulations, or additional strategies to generate antibodies that neutralize a full spectrum of transmitted HIV-1 strains.

In most cases of HIV-1 transmission, only a subset of variants is transmitted from the index case to the newly infected individual. Understanding the characteristics of these transmitted variants may aid in developing new methods to halt the spread of HIV-1. Studies evaluating the genotypic and antigenic properties of transmitted variants have provided insights into how the selective pressures applied during different modes of transmission uniquely imprint the infecting viruses. In the setting of sexual transmission, variants with increased neutralization sensitivity appeared to be selected during transmission in discordant subtype C-infected couples, although transmitted variants from different risk groups and HIV-1 subtypes did not demonstrate increased neutralization sensitivity, suggesting this may not be a consistent feature of transmitted variants. Studies of both mother to child transmission (MTCT) and superinfection, where preexisting NAbs are present at the time of exposure, provide opportunities to analyze whether the breadth and potency of the NAb response influence the incidence of new infections. MTCT resulted in selection for variants that were resistant to maternal antibodies, suggesting that maternal antibodies can protect the baby from those variants that are susceptible to the antibodies present. There are some data to suggest that poor neutralizing antibody (NAb) responses are present in cases of superinfection, although these data are preliminary. Defining the characteristics of the viruses transmitted in the presence and absence of NAbs as well as defining the NAb responses that fail to protect from infection during MTCT and superinfection may provide critical insights into the antibody responses that are needed for effective vaccines and other prophylactic therapeutics.

The mucosal immune response to HIV weighs in heavily on the battle against it, as the majority of infections occur via the mucosal route. The antibody response in the mucosae, specifically the genital tract, is characterized by binding and, in some studies, neutralizing HIV-specific IgG and IgA antibodies. Ample evidence, however, points to discrepancies and difficulties in the detection of HIV-specific IgA in HIV-positive subjects, and an even more pronounced divide surfaces in studies done with individuals exposed to HIV, but uninfected. Reports in the literature detail HIV-specific (in some cases, neutralizing) IgA antibodies, in the absence of specific IgG, in the serum and mucosal secretions of virusexposed, seronegative subjects; this has given rise to speculation that HIV-specific IgA provides a protective immune response to the virus in high-risk individuals who remain seronegative. Contradictory results, however, describe the absence of both IgA and IgG HIV antibodies in the mucosal secretions of similar cohorts. Considering the importance of the antibody response to ascertaining the correlates of HIV immunity, as well as on vaccine research and development, this review addresses the relevant studies and their implications.

Over the last two decades, use of SIV for experimental infection of Asian macaques has provided important leads in the quest for an AIDS vaccine, served as the genesis of recombinant SIV/HIV viruses (SHIV), and perhaps most importantly, helped establish or confirm biological relevance for a variety of hypotheses related to the host immune response to infection and the corresponding viral strategies for evading that response. The env genes of HIV-1 and SIV encode proteins bearing a high degree of structural similarity and sharing an identical suite of essential functions. The Env complexes formed by these proteins are present on the surface of virus-producing cells and virions, where they are the primary targets of the host neutralizing antibody response. In this review, we briefly describe the similarities between HIV-1 infection and SIV experimental models, then focus specifically on the use of the SIV/macaque model as a tool for understanding the humoral immune response to infection and resistance to antibody-mediated neutralization in HIV infection and AIDS.

Antibody (Ab) mediated neutralization is a crucial means of host resistance to many pathogens and will most likely be required in the development of a vaccine to protect against HIV-1. Here we examine mechanistic aspects of HIV-1 neutralization with attention to recent studies on the stoichiometric, kinetic and thermodynamic parameters involved. Neutralization of HIV-1, as with any microbe, minimally requires an initial molecular encounter with Ab. Ab occupancy of functional heterotrimers of the envelope glycoproteins, gp120 and gp41 (Env), indeed appears to be the dominant mechanism of neutralization for HIV-1. However, the Ab-binding site, the parameters mentioned above, as well as the stages and duration of vulnerability to Ab recognition, prior to and leading up to viral entry, each have a distinct impact on the mechanism of neutralization for any given Ab specificity. With HIV-1, the problems of mutational variation and neutralization resistance, coupled with the lability and conformational heterogeneity in Env, have stimulated the search for rational approaches to Env immunogen design that are unprecedented in vaccinology.

Upon viral exposure, antibodies serve as a first line of defense and can act by preventing infection or reducing the viral burden. The ability of antibodies to confer protection against HIV has been demonstrated by several studies using the passive transfer of neutralizing antibodies in the non-human primate challenge model. Therefore, efforts have been made to induce a similarly protective humoral immune response by vaccination with antigens derived from HIV. Thus far, the results have been disappointing. Humoral immune responses elicited via vaccination display activities that are generally much less potent and broad as compared to those induced during natural infection. However, recently there have been increased efforts to systematically identify and compare the epitopes potentially critical to the generation of protective antibody responses in the hope that this will lead to improved strategies and superior immunogen design. As a critical part of this process, novel methods to monitor protective antibody responses will also need to be vigorously explored and improved, then validated in both preclinical and clinical settings.

The HIV-1/AIDS epidemic continues to escalate, and a protective vaccine remains elusive. The first vaccine candidate, gp120, did not induce broadly neutralizing antibodies (nAbs) against primary HIV-1 isolates and was ineffective in phase III clinical trials. Attention then focused on generating cytotoxic lymphocyte (CTL)-based vaccines. Interest in anti-HIV-1 nAbs was renewed when passive immunization with human neutralizing monoclonal antibodies (nmAbs) completely protected macaques after intravenous and mucosal challenges with simian-human immunodeficiency viruses (SHIVs) encoding HIV-1 env. These nmAbs targeted conserved, functionally important epitopes on gp120 and gp41. Protection in primate/SHIV models was observed when nmAbs were used singly (nmAbs 2G12, b12) and in various combination regimens (nmAbs b12, F105, 2G12, 2F5, 4E10). Passive immunization, a well-established tool to determine the correlates of protective immunity, thus identified protective epitopes. The three-dimensional structures of some of the latter were recently elucidated, generating important information to design nAb-response-base immunogens. However, several of the protective nmAbs were found to exhibit autoreactivity, raising the possibility that B-cell responses against the cognate epitopes may be difficult to induce by active immunization. It will be important to explore whether broad neutralization can be dissociated from autoreactivity. Future experiments will reveal whether other conserved HIV-1 Env epitopes exist, antibodies against which will be broadly neutralizing in vitro, protective as passive immunization in SHIVchallenged macaques, but lacking autoreactivity. Since all protective epitopes identified to date are located on HIV-1 clade B Env, future studies should include analysis of nmAbs against non-clade B strains.