Elsevier

Vaccine

Volume 24, Issue 26, 29 June 2006, Pages 5442-5451
Vaccine

HIV-1 Envgp140 trimers elicit neutralizing antibodies without efficient induction of conformational antibodies

https://doi.org/10.1016/j.vaccine.2006.03.063Get rights and content

Abstract

Currently, no vaccine for human immunodeficiency virus (HIV-1) provides protection from virus infection. One reason for these disappointing results has been the difficulty of current vaccine candidates to elicit high-titer, broadly reactive immunity to a large number of viral proteins. Recently, our laboratory demonstrated that the coupling of C3d to a soluble trimerized HIV-1 envelope (Envgp140(FT)) elicited higher titers of neutralizing antibodies than monomers of Envgp120 coupled to C3d [Bower JF, Yang X, Sodroski J, Ross TM. Elicitation of neutralizing antibodies with DNA vaccines expressing soluble stabilized human immunodeficiency virus type 1 envelope glycoprotein trimers conjugated to C3d. J Virol 2004;78(9):4710–9]. To determine if the induction of conformational antibodies correlated with neutralization, mice (BALB/c) were primed (2×) with DNA plasmids expressing monomeric Envgp120 or trimeric Envgp140 alone or fused to mC3d3 at one of two doses (2.0 μg or 0.2 μg), followed by a boost of recombinant uncleaved, trimeric Envgp140. Regardless of the priming dose of DNA, all mice had high-titer anti-Env IgG antibodies. Interestingly, Envgp140 trimers did not elicit higher titers of antibodies that recognized conformational Env epitopes compared to monomers of Envgp120. Therefore, additional parameters were examined for correlation with neutralization. For neutralization-resistant HIV-1 isolates, ADA and YU-2, neutralization correlated with high-titer, high avidity antibodies, with Envgp140 eliciting slightly higher neutralization titers than Envgp120. In contrast, none of the measured parameters correlated with neutralization for the more neutralization-sensitive isolates, MN or 89.6. Therefore, even though soluble, uncleaved Envgp140 trimers may be marginally more effective at eliciting neutralizing antibodies than Envgp120, neutralization does not appear to correlate with the elicitation of conformationally dependent antibodies.

Introduction

Despite a variety of vaccine approaches that elicit diverse immune responses, currently no HIV/AIDS vaccine is effective at preventing viral infection [1], [2], [3], [4], [5], [6], [7], [8], [9]. A lack of understanding of the immune correlates for protection is a primary reason for the ineffectiveness of past AIDS vaccines [10]. Previous passive immunization studies demonstrated that sterilizing immunity against simian immunodeficiency virus (SIV) and simian-human immunodeficiency virus (SHIV) infection is possible [11], [12], [13], [14]. However, a successful vaccine against HIV-1 should not only have the capacity to induce HIV-1 specific antibodies, but also to potent cellular immunity to clear virally infected cells [15], [16], [17].

Previous HIV-1/AIDS vaccines utilizing live-attenuated virus, viral vectors, and recombinant subunit proteins have been tested with varying degrees of success (for reviews please see [18], [19], [20], [21]). In the past decade, vaccination with DNA plasmids has offered an alternative strategy for the elicitation of both humoral and cellular immune responses without some of the safety issues associated with live virus or viral vectors [22], [23], [24], [25]. However, the efficient elicitation of immune responses by DNA vaccines in small rodents has not translated into the elicitation of high-titer, protective immunity in non-human primates [16], [26], [27]. Priming the immune system with a DNA plasmid(s), followed by one of a variety of booster inoculations (i.e. viral vectors or recombinant proteins), has proven an effective strategy for eliciting high-titer immune responses to HIV-1 antigens [16], [26].

The role of Env appears critical for the induction of protective immunity in recent AIDS vaccine candidates tested in non-human primates [28], [29]. Inclusion of Env in a multi-component AIDS vaccine resulted in lower viral set points and higher CD4 counts following challenge compared to the same vaccines lacking Env. Envelope on the native virion most likely forms a trimer, although one model suggests that both functional and non-functional trimeric spikes are present on virions. Several vaccine strategies have incorporated an oligomeric/trimeric form of Env in order to elicit cross-reactive immunity that neutralizes viral infection [1], [4], [5], [8], [30], [31], [32], [33], [34], [35], [36], [37], [38]. Soluble, stabilized Env trimers are designed to better mimic the trimeric structure of the envelope on the native virion compared to monomeric immunogens. The goal of trimer immunogen design is to present and preferentially induce conformationally dependent antibodies that recognize epitopes present only on the native virion-associated spikes. Several of these trimeric Env immunogens do elicit significantly slightly higher titers of neutralizing antibodies than monomeric Envgp120 [32], [38], [39], [40], however the breadth of neutralization is still somewhat limited. Often, these oligomeric Env proteins are produced by eliminating the natural cleavage site recognized by cellular proteases [37], [41], [42], [43], which might influence the trimeric structure [4], [5]. The lack of elicited high-titer, broadly reactive neutralizing antibodies by these immunogens may be associated with the elicitation of primarily non-neutralizing antibodies [5], [44], [45], which may because these uncleaved envelopes (1) are in non-native forms or (2) are processed through different cellular pathways than cleaved forms of Env [4], [5], [46], [47].

In this study, soluble, stabilized trimeric Envgp140 molecules were constructed and used as immunogens in a DNA prime/trimerized Envgp140 protein boost regimen to examine if the induced immunity potentially correlates with neutralization. Each trimeric Envgp140 molecule contained the Envgp120 exterior of Env and the ectodomain of Envgp41, stabilized with synthetic trimerization motifs from the T4 bacteriophage fibritin (FT) [Envgp140(FT)] [37]. This motif appears as stable in vitro as the previously described eukaryotic GCN4 stabilization motif [Envgp140(GCN4)] [36], [37]. Even though uncleaved Envgp140 trimers are not yet optimally designed, these proteins did elicit marginally higher neutralizing titers than monomers of Envgp120 [32] and therefore, in this study the goal was to determine whether these elicited neutralizing antibodies correlated with the induction of conformationally dependent antibodies.

Section snippets

Plasmid DNA

The pTR600 eukaryotic expression vector, the monomeric, soluble Envgp120 or the FT stabilized Envgp140 from the isolate, YU-2 (accession # M93258), have been previously described [32]. Briefly, the plasmids expressing Envgp120 contain a gene segment expressing the entire ectodomain of gp120 (nt 1–1500). The Envgp140 plasmid expresses a protein representing the entire ectodomains from gp120 and gp41, which was stabilized by the addition of the synthetic trimerization domain derived from the T4

Elicitation of anti-Env antibodies in vaccinated mice

Mice (BALB/c) were vaccinated via gene gun on Day 1 and week 4 with either high dose DNA (2.0 μg) or low dose DNA (0.2 μg) plasmids. Regardless if the mice were primed with monomeric Envgp120 or trimeric Envgp140 or if the envelopes were conjugated to mC3d3, each mouse had anti-Env IgG titers after the second DNA plasmid inoculation (week 6), which rose following a boost of recombinant Envgp140 (Fig. 2). Mice vaccinated with a high dose of DNA expressing any of the Env immunogens had anti-Env

Discussion

In this study, several vaccine strategies were combined to elicit anti-Env immune responses. Mice were vaccinated with two doses of DNA plasmid expressing soluble forms of Env followed by a booster inoculation of a purified, recombinant, trimerized Envgp140 protein. Mice were primed with either a high dose of DNA (2.0 μg) or a low dose of DNA (0.2 μg), expressing one of two forms of envelope (monomeric gp120 or trimeric gp140). Additional groups of mice were vaccinated with DNA expressing

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    Present address: Global Clinical Immunology, Sanofi Pasteur, Discovery Drive, Swiftwater, PA 18370, USA. Tel.: +1 570 895 3225; fax: +1 570 895 2872.

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