Comparative Immunogenicity of HIV-1 gp160, gp140 and gp120 Expressed by Live Attenuated Newcastle Disease Virus Vector

The development of a vaccine against human immunodeficiency virus-1 (HIV-1) capable of inducing broad humoral and cellular responses at both the systemic and mucosal levels will be critical for combating the global AIDS epidemic. We previously demonstrated the ability of Newcastle disease virus (NDV) as a vaccine vector to express oligomeric Env protein gp160 and induce potent humoral and mucosal immune responses. In the present study, we used NDV vaccine strain LaSota as a vector to compare the biochemical and immunogenic properties of vector-expressed gp160, gp120, and two versions of gp140 (a derivative of gp160 made by deleting the transmembrane and cytoplasmic domains), namely: gp140L, which contained the complete membrane-proximal external region (MPER), and gp140S, which lacks the distal half of MPER. We show that, similar to gp160, NDV-expressed gp140S and gp120, but not gp140L, formed higher-order oligomers that retained recognition by conformationally sensitive monoclonal antibodies. Immunization of guinea pigs by the intranasal route with rLaSota/gp140S resulted in significantly greater systemic and mucosal antibody responses compared to the other recombinants. Immunization with rLaSota/140S, rLaSota/140L rLaSota/120 resulted in mixed Th1/Th2 immune responses as compared to Th1-biased immune responses induced by rLaSota/160. Importantly, rLaSota/gp140S induced neutralizing antibody responses to homologous HIV-1 strain BaL.26 and laboratory adapted HIV-1 strain MN.3 that were stronger than those elicited by the other NDV recombinants. Additionally, rLaSota/gp140S induced greater CD4+ and CD8+ T-cell responses in mice. These studies illustrate that rLaSota/gp140S is a promising vaccine candidate to elicit potent mucosal, humoral and cellular immune responses to the HIV-1 Env protein.     

Intranasal vaccination using interleukin-12 and cholera toxin subunit B as adjuvants to enhance mucosal and systemic immunity to human immunodeficiency virus type 1 glycoproteins

We have investigated the induction of protective mucosal immunity to human immunodeficiency virus type 1 (HIV-1) isolate 89.6 by intranasal (i.n.) immunization of mice with gp120 and gp140 together with interleukin-12 (IL-12) and cholera toxin subunit B (CTB) as adjuvants. It was found that both IL-12 and CTB were required to elicit mucosal antibody responses and that i.n. immunization resulted in increased total, immunoglobulin G1 (IgG1), and IgG2a anti-HIV-1 antibody levels in serum; increased total, IgG1, IgG2a, and IgA antibody expression in bronchoalveolar lavage fluids; and increased IgA antibody levels in vaginal washes. Levels of anti-HIV-1 antibodies in both sera and secretions were higher in groups immunized with gp140 than in those immunized with gp120. However, only gp120-specific mucosal antibodies demonstrated neutralizing activity against HIV-1 89.6. Taken together, the results show that IL-12 and CTB act synergistically to enhance both systemic and local mucosal antibody responses to HIV-1 glycoproteins and that even though gp140 induces higher antibody titers than gp120, only gp120-specific mucosal antibodies interfere with virus infectivity.     

Enhanced Immune Responses to HIV-1 Envelope Elicited by a Vaccine Regimen Consisting of Priming with Newcastle Disease Virus Expressing HIV gp160 and Boosting with gp120 and SOSIP gp140 Proteins

Newcastle disease virus (NDV) expressing HIV-1 BaL gp160 was evaluated either alone or with monomeric BaL gp120 and BaL SOSIP gp140 protein in a prime-boost combination in guinea pigs to enhance envelope (Env)-specific humoral and mucosal immune responses. We showed that a regimen consisting of an NDV prime followed by a protein boost elicited stronger serum and mucosal Th-1-biased IgG responses and neutralizing antibody responses than NDV-only immunizations. Additionally, these responses were higher after the gp120 than after the SOSIP gp140 protein boost.     

Systemic immunization with an ALVAC-HIV-1/protein boost vaccine strategy protects rhesus macaques from CD4+ T-cell loss and reduces both systemic and mucosal simian-human immunodeficiency virus SHIVKU2 RNA levels

Transmission of human immunodeficiency virus type 1 (HIV-1) occurs primarily via the mucosal route, suggesting that HIV-1 vaccines may need to elicit mucosal immune responses. Here, we investigated the immunogenicity and relative efficacy of systemic immunization with two human ALVAC-HIV-1 recombinant vaccines expressing Gag, Pol, and gp120 (vCP250) or Gag, Pol, and gp160 (vCP1420) in a prime-boost protocol with their homologous vaccine native Env proteins. The relative efficacy was measured against a high-dose mucosal exposure to the pathogenic neutralization-resistant variant SHIV(KU2) (simian-human immunodeficiency virus). Systemic immunization with both vaccine regimens decreased viral load levels not only in blood but unexpectedly also in mucosal sites and protected macaques from peripheral CD4+ T-cell loss. This protective effect was stronger when the gp120 antigen was included in the vaccine. Inclusion of recombinant Tat protein in the boosting phase along with the Env protein did not contribute further to the preservation of CD4+ T cells. Thus, systemic immunization with ALVAC-HIV-1 vaccine candidates elicits anti-HIV-1 immune responses able to contain virus replication also at mucosal sites in macaques.     

Antibodies with specificity to native gp120 and neutralization activity against primary human immunodeficiency virus type 1 isolates elicited by immunization with oligomeric gp160.

Current human immunodeficiency virus type 1 (HIV-1) envelope vaccine candidates elicit high antibody binding titers with neutralizing activity against T-cell line-adapted but not primary HIV-1 isolates. Serum antibodies from these human vaccine recipients were also found to be preferentially directed to linear epitopes within gp120 that are poorly exposed on native gp120. Systemic immunization of rabbits with an affinity-purified oligomeric gp160 protein formulated with either Alhydrogel or monophosphoryl lipid A-containing adjuvants resulted in the induction of high-titered serum antibodies that preferentially bound epitopes exposed on native forms of gp120 and gp160, recognized a restricted number of linear epitopes, efficiently bound heterologous strains of monomeric gp120 and cell surface-expressed oligomeric gp120/gp41, and neutralized several strains of T-cell line-adapted HIV-1. Additionally, those immune sera with the highest oligomeric gp160 antibody binding titers had neutralizing activity against some primary HIV-1 isolates, using phytohemagglutinin-stimulated peripheral blood mononuclear cell targets. Induction of an antibody response preferentially reactive with natively folded gp120/gp160 was dependent on the tertiary structure of the HIV-1 envelope immunogen as well as its adjuvant formulation, route of administration, and number of immunizations administered. These studies demonstrate the capacity of a soluble HIV-1 envelope glycoprotein vaccine to elicit an antibody response capable of neutralizing primary HIV-1 isolates.     

Induction of a Mucosal Cytotoxic T-Lymphocyte Response by Intrarectal Immunization with a Replication-Deficient Recombinant Vaccinia Virus Expressing Human Immunodeficiency Virus 89.6 Envelope Protein

To improve the safety of recombinant vaccinia virus vaccines, modified vaccinia virus Ankara (MVA) has been employed, because it has a replication defect in most mammalian cells. Here we apply MVA to human immunodeficiency virus type 1 (HIV-1) vaccine development by incorporating the envelope protein gp160 of HIV-1 primary isolate strain 89.6 (MVA 89.6) and use it to induce mucosal cytotoxic-T-lymphocyte (CTL) immunity. In initial studies to define a dominant CTL epitope for HIV-1 89.6 gp160, we mapped the epitope to a sequence, IGPGRAFYAR (from the V3 loop), homologous to that recognized by HIV MN loop-specific CTL and showed that HIV-1 MN-specific CTLs cross-reactively recognize the corresponding epitope from strain 89.6 presented by H-2D^d. Having defined the CTL specificity, we immunized BALB/c mice intrarectally with recombinant MVA 89.6. A single mucosal immunization with MVA 89.6 was able to elicit long-lasting antigen-specific mucosal (Peyer’s patch and lamina propria) and systemic (spleen) CTL responses as effective as or more effective than those of a replication-competent vaccinia virus expressing 89.6 gp160. Immunization with MVA 89.6 led to (i) the loading of antigen-presenting cells in vivo, as measured by the ex vivo active presentation of the P18-89.6 peptide to an antigen-specific CTL line, and (ii) the significant production of the proinflammatory cytokines (interleukin-6 and tumor necrosis factor alpha) in the mucosal sites. These results indicate that nonreplicating recombinant MVA may be at least as effective for mucosal immunization as replicating recombinant vaccinia virus.     

Mucosal and systemic immune responses to a human immunodeficiency virus type 1 epitope induced upon vaginal infection with a recombinant influenza A virus

The humoral and cellular immune responses in the genital mucosa likely play an important role in the prevention of sexually transmitted infections, including infection with human immunodeficiency virus type 1 (HIV-1). Here we show that vaginal infection of progesterone-treated BALB/c mice with a recombinant influenza virus bearing the immunodominant P18IIIB cytotoxic T-lymphocyte (CTL) epitope of the gp160 envelope protein from an HIV-1 IIIB isolate (P18IIIB; RIQRGPGRAFVTIGK) can induce a specific immune response in regional mucosal lymph nodes, as well as in a systemic site (the spleen). A single inoculation of mice with the recombinant influenza virus induced long-lasting (at least 5 months) antigen-specific CTL memory detectable as a rapid recall of effector CTLs upon vaginal infection with recombinant vaccinia virus expressing HIV-1 IIIB envelope gene products. Long-term antigen-specific CTL memory was also induced and maintained in distant mucosal tissues when mice were intranasally immunized with the recombinant influenza virus. These results indicate that mucosal immunization and, in particular, local vaginal immunization with recombinant influenza virus can provide strong, durable immune responses in the female genital tract of mice.     

Evaluation of genetic immunization adjuvants to improve the effectiveness of a human immunodeficiency virus type 2 (HIV-2) envelope DNA vaccine

In an effort to develop a more effective genetic immunization strategy for HIV, we developed an HIV-2 env DNA vaccine and evaluated three adjuvant formulations. The gp140 gene from HIV-2(UC2 )was synthesized using mammalian codons and cloned into a plasmid vector that expresses eukaryotic genes at high levels. We found that after three immunizations in mice, a novel cationic liposome formulation (Vaxfectin) was superior at inducing systemic and mucosal antibody responses compared to a naked DNA, a controlled release device (an Alzet minipump) and polysaccharide microparticles made from chitosan (P = 0.027). Vaxfectin also induced higher levels of systemic antibodies for each isotype and IgG subclass as well as levels of HIV-2-specific mucosal IgA (P = 0.034). When different routes of immunization were used with the Vaxfectin formulation, gp140-specific systemic antibody responses were highest by the intradermal route, mucosal antibody responses were highest by the intramuscular route, while the intranasal route was the least effective. These results suggest that this cationic liposome formulation is an important adjuvant to improve the effectiveness of genetic immunization strategies for AIDS, and that multiple routes of immunization should be employed for optimal efficacy for HIV vaccine candidates.     

A recombinant Newcastle disease virus (NDV) expressing VP2 protein of infectious bursal disease virus (IBDV) protects against NDV and IBDV

Infectious bursal disease virus (IBDV) causes a highly immunosuppressive disease in chickens. Currently available, live IBDV vaccines can lead to generation of variant viruses. We have developed an alternative vaccine that will not create variant IBDV. By using the reverse genetics approach, we devised a recombinant Newcastle disease virus (NDV) vector from a commonly used vaccine strain LaSota to express the host-protective immunogen VP2 of a variant IBDV strain GLS-5. The gene encoding the VP2 protein of the IBDV was inserted into the most 3'-proximal locus of a full-length NDV cDNA for high-level expression. We successfully recovered the recombinant virus, rLaSota/VP2. The rLaSota/VP2 was genetically stable, at least up to 12 serial passages in chicken embryos, and was shown to express the VP2 protein. The VP2 protein was not incorporated into the virions of recombinant virus. Recombinant rLaSota/VP2 replicated to a titer similar to that of parental NDV strain LaSota in chicken embryos and cell cultures. To assess protective efficacy of the rLaSota/VP2, 2-day-old specific-pathogen-free chickens were vaccinated with the recombinant virus and challenged with a highly virulent NDV strain Texas GB or IBDV variant strain GLS-5 at 3 weeks postvaccination. Vaccination with rLaSota/VP2 generated antibody responses against both NDV and IBDV and provided 90% protection against NDV and IBDV. Booster immunization induced higher levels of antibody responses against both NDV and IBDV and conferred complete protection against both viruses. These results indicate that the recombinant NDV can be used as a vaccine vector for other avian pathogens.     

Protective immunity induced by systemic and mucosal delivery of DNA vaccine expressing glycoprotein B of pseudorabies virus

A murine model immunized by systemic and mucosal delivery of plasmid DNA vaccine expressing glycoprotein B (pCIgB) of pseudorabies virus (PrV) was used to evaluate both the nature of the induced immunity and protection against a virulent virus. With regard to systemic delivery, the intramuscular (i.m.) immunization with pCIgB induced strong PrV-specific IgG responses in serum but was inefficient in generating a mucosal IgA response. Mucosal delivery through intranasal (i.n.) immunization of pCIgB induced both systemic and mucosal immunity at the distal mucosal site. However, the levels of systemic immunity induced by i.n. immunization were less than those induced by i.m. immunization. Moreover, i.n. genetic transfer of pCIgB appeared to induce Th2-biased immunity compared with systemic delivery, as judged by the ratio of PrV-specific IgG isotypes and Th1- and Th2-type cytokines produced by stimulated T cells. Moreover, the immunity induced by i.n. immunization did not provide effective protection against i.n. challenge of a virulent PrV strain, whereas i.m. immunization produced resistance to viral infection. Therefore, although i.n. immunization was a useful route for inducing mucosal immunity at the virus entry site, i.n. immunization did not provide effective protection against the lethal infection of PrV.     

Characterization of the outer domain of the gp120 glycoprotein from human immunodeficiency virus type 1

The core of the gp120 glycoprotein from human immunodeficiency virus type 1 (HIV-1) is comprised of three major structural domains: the outer domain, the inner domain, and the bridging sheet. The outer domain is exposed on the HIV-1 envelope glycoprotein trimer and contains binding surfaces for neutralizing antibodies such as 2G12, immunoglobulin G1b12, and anti-V3 antibodies. We expressed the outer domain of HIV-1(YU2) gp120 as an independent protein, termed OD1. OD1 efficiently bound 2G12 and a large number of anti-V3 antibodies, indicating its structural integrity. Immunochemical studies with OD1 indicated that antibody responses against the outer domain of the HIV-1 gp120 envelope glycoprotein are rare in HIV-1-infected human sera that potently neutralize the virus. Surprisingly, such outer-domain-directed antibody responses are commonly elicited by immunization with recombinant monomeric gp120. Immunization with soluble, stabilized HIV-1 envelope glycoprotein trimers elicited antibody responses that more closely resembled those in the sera of HIV-1-infected individuals. These results underscore the qualitatively different humoral immune responses elicited during natural infection and after gp120 vaccination and help to explain the failure of gp120 as an effective vaccine.     

Prime-boost immunization schedules based on influenza virus and vaccinia virus vectors potentiate cellular immune responses against human immunodeficiency virus Env protein systemically and in the genitorectal draining lymph nodes

Vaccines that elicit systemic and mucosal immune responses should be the choice to control human immunodeficiency virus (HIV) infections. We have previously shown that prime-boost immunizations with influenza virus Env and vaccinia virus (VV) WR Env recombinants induced an enhanced systemic CD8(+) T-cell response against HIV-1 Env antigen. In this report, we analyzed in BALB/c mice after priming with influenza virus Env the ability of two VV recombinants expressing HIV-1 Env B (VV WR Env and the highly attenuated modified VV Ankara [MVA] Env) to boost cellular immune responses in the spleen and in the lymph nodes draining the genital and rectal tracts. Groups of mice were primed by the intranasal route with 10(4) PFU of influenza virus Env and boosted 14 days later by the intraperitoneal or intranasal route with 10(7) PFU of MVA Env or VV WR Env, while the control group received two immunizations with influenza virus Env. We found that the combined immunization (Flu/VV) increased more than 60 times the number of gamma interferon-specific CD8(+) T cells compared to the Flu/Flu scheme. Significantly, boosting with MVA Env by the intraperitoneal route induced a response 1.25 or 2.5 times (spleen or genital lymph nodes) higher with respect to that found after the boost with VV WR Env. Mice with an enhanced CD8(+) T-cell response also had an increased Th1/Th2 ratio, evaluated by the cytokine pattern secreted following in vitro restimulation with gp160 protein and by the specific immunoglobulin G2a (IgG2a)/IgG1 ratio in serum. By the intranasal route recombinant WR Env booster gave a more efficient immune response (10 and 1.3 times in spleen and genital lymph nodes, respectively) than recombinant MVA Env. However, the scheme influenza virus Env/MVA Env increased four times the response in the spleen, giving a low but significant response in the genital lymph nodes compared with a single intranasal immunization with MVA Env. These results demonstrate that the combination Flu/MVA in prime-booster immunization regimens is an effective vaccination approach to generate cellular immune responses to HIV antigens at sites critical for protective responses.     

Biochemical and immunogenic characterization of soluble human immunodeficiency virus type 1 envelope glycoprotein trimers expressed by semliki forest virus

The current lack of envelope glycoprotein immunogens that elicit broadly neutralizing antibody responses remains a major challenge for human immunodeficiency virus type 1 (HIV-1) vaccine development. However, the recent design and construction of stable soluble gp140 trimers have shown that some neutralization breadth can be achieved by using immunogens that better mimic the functional viral spike complex. The use of genetic delivery systems to drive the in vivo expression of such immunogens for the stimulation of neutralizing antibodies against HIV-1 may offer advantages by maintaining the quaternary structure of the trimeric envelope glycoproteins. Here, we describe the biochemical and immunogenic properties of soluble HIV-1 envelope glycoprotein trimers expressed by recombinant Semliki Forest virus (rSFV). The results presented here demonstrate that rSFV supports the expression of stable soluble gp140 trimers that retain recognition by conformationally sensitive antibodies. Further, we show that rSFV particle immunizations efficiently primed immune responses as measured after a single boost with purified trimeric gp140 protein, resulting in a Th1-biased antibody response. This differed from the Th2-biased antibody response obtained after repeated immunizations with purified gp140 protein trimers. Despite this difference, both regimens stimulated neutralizing antibody responses of similar potency. This suggests that rSFV may be a useful component of a viral vector prime-protein boost regimen aimed at stimulating both cell-mediated immune responses and neutralizing antibodies against HIV-1.     

Common Features of Mucosal and Peripheral Antibody Responses Elicited by Candidate HIV-1 Vaccines in Rhesus Monkeys

Human immunodeficiency virus type 1 (HIV-1) vaccines that elicit protective antibody responses at mucosal sites would be highly desirable. Here, we report that intramuscular immunization of candidate HIV-1 vaccine vectors and purified Env proteins elicited potent and durable humoral immune responses in colorectal mucosa in rhesus monkeys. The kinetics, isotypes, functionality, and epitope specificity of these mucosal antibody responses were similar to those of peripheral responses in serum. These data suggest a close immunological relationship between mucosal and systemic antibody responses following vaccination in primates.     

Transmembrane envelope glycoproteins of human immunodeficiency virus type 2 and simian immunodeficiency virus SIV-mac exist as homodimers.

An 80-kilodalton glycoprotein (gp80) was produced in human immunodeficiency virus type 2 (HIV-2)-infected cells along with three envelope glycoproteins that we have recently reported: the extracellular glycoprotein (gp125), the envelope glycoprotein precursor (gp140), and the transient dimeric form of the precursor (gp300). gp125 and gp80 were detectable after the synthesis of gp140 and the formation of gp300. Using a specific monoclonal antibody, we showed here that gp80 is a dimeric form of the transmembrane glycoprotein gp36 of HIV-2. Dimerization of the envelope glycoprotein precursor and dimeric forms of the transmembrane glycoproteins were also observed in cells infected with simian immunodeficiency virus (SIV-mac), a virus closely related to HIV-2. Under routine conditions of our experiments (i.e., extraction by 1% Triton X-100 before polyacrylamide gel electrophoresis in sodium dodecyl sulfate [SDS]), monomeric forms of the transmembrane glycoprotein of HIV-2 and SIV-mac were only seldomly observed. Dimeric forms of the envelope precursors and the transmembrane glycoproteins are probably stabilized by extraction in the nonionic detergent Triton X-100 since such dimeric forms resist dissociation during subsequent electrophoresis in the presence of the ionic detergent SDS. However, the dissociation of these dimeric forms might occur when samples are prepared by extraction directly in 1% SDS or by incubation of the purified dimers at acidic pH. Dimerization of the envelope precursor might be required for its processing to give the mature envelope proteins, whereas the transmembrane dimer might be essential for optimal structure of the virion and thus its infectivity.     

Solid-phase proteoliposomes containing human immunodeficiency virus envelope glycoproteins

The human immunodeficiency virus type 1 (HIV-1) exterior envelope glycoprotein gp120 mediates receptor binding and is the major target for neutralizing antibodies. A broadly neutralizing antibody response is likely to be a critical component of the immune response against HIV-1. Although antibodies against monomeric gp120 are readily elicited in immunized individuals, these antibodies are inefficient in neutralizing primary HIV-1 isolates. As a chronic pathogen, HIV-1 has evolved to avoid an optimal host response by a number of immune escape mechanisms. Monomeric gp120 that has dissociated from the functional trimer presents irrelevant epitopes that are not accessible on functional trimeric envelope glycoproteins. The resulting low level of antigenic cross-reactivity between monomeric gp120 and the functional spike may contribute to the inability of monomeric gp120 to elicit broadly neutralizing antibodies. Attempts to generate native, trimeric envelope glycoproteins as immunogens have been frustrated by both the lability of the gp120-gp41 interaction and the weak association between gp120 subunits. Here, we present solid-phase HIV-1 gp160DeltaCT (cytoplasmic tail-deleted) proteoliposomes (PLs) containing native, trimeric envelope glycoproteins in a physiologic membrane setting. We present data that indicate that the gp160DeltaCT glycoproteins on PLs are trimers and are recognized by several relevant conformational ligands in a manner similar to that for gp160DeltaCT oligomers expressed on the cell surface. The PLs represent a significant advance over present envelope glycoprotein formulations as candidate immunogens for HIV vaccine design and development.     

Human immunodeficiency virus type 1 neutralization by a single molecule of V3-targeted antibody

Human immunodeficiency virus type-1 (HIV-1) infection generally provokes antibody responses to the viral envelope glycoprotein. Two major regions of gp120, the third variable (V3) domain and the CD4-binding site, have been identified as neutralization targets. The precise mechanism of HIV-1 neutralization by antibodies against the V3 domain is still unknown. It is shown that by kinetic neutralization studies, one molecule of V3-targeted monoclonal antibody (0.5beta) is enough to neutralize one virion. This antibody, which neutralized more than 99% of the virus, inhibited the binding of the virus to cells by 42%. HIV-1 pseudotyped with G glycoprotein from vesicular stomatitis virus was also neutralized by 0.5beta, suggesting that the antibody did not inhibit the viral attachment but caused some alteration in the envelope. These results indicate that the antibody plays an additional role on steric change of the envelope involved in inhibition of viral entry.     

Rabies virus-based vectors expressing human immunodeficiency virus type 1 (HIV-1) envelope protein induce a strong, cross-reactive cytotoxic T-lymphocyte response against envelope proteins from different HIV-1 isolates

Novel viral vectors that are able to induce both strong and long-lasting immune responses may be required as effective vaccines for human immunodeficiency virus type 1 (HIV-1) infection. Our previous experiments with a replication-competent vaccine strain-based rabies virus (RV) expressing HIV-1 envelope protein from a laboratory-adapted HIV-1 strain (NL4-3) and a primary HIV-1 isolate (89.6) showed that RV-based vectors are excellent for B-cell priming. Here we report that cytotoxic T-lymphocyte (CTL) responses against HIV-1 gp160 are induced by recombinant RVs. Our results indicated that a single inoculation of mice with an RV expressing HIV-1 gp160 induced a solid and long-lasting memory CTL response specific for HIV-1 envelope protein. Moreover, CTLs from immunized mice were not restricted to the homologous HIV-1 envelope protein and were able to cross-kill target cells expressing HIV-1 gp160 from heterologous HIV-1 strains. These studies further suggest promise for RV-based vectors to elicit a persistent immune response against HIV-1 and their potential utility as efficacious anti-HIV-1 vaccines.     

Induction of systemic and mucosal cross-clade neutralizing antibodies in BALB/c mice immunized with human immunodeficiency virus type 1 clade A virus-like particles administered by different routes of inoculation

We have recently developed a candidate human immunodeficiency virus type 1 (HIV-1) vaccine model, based on virus-like particles (VLPs) expressing gp120 from a Ugandan HIV-1 isolate of clade A (HIV-VLP(A)s), which shows the induction of neutralizing antibodies as well as cytotoxic T lymphocytes (CTL) in BALB/c mice by intraperitoneal (i.p.) administration. In the present study, immunization experiments based on a multiple-dose regimen have been performed with BALB/c mice to compare different routes of administration. i.p. and intranasal (i.n.), but not oral, administration induce systemic as well as mucosal (vaginal and intestinal) immunoglobulin G (IgG) and IgA responses. These immune sera exhibit >50% ex vivo neutralizing activity against both autologous and heterologous primary isolates. Furthermore, the administration of HIV-VLP(A)s by the i.n. immunization route induces a specific CTL activity, although at lower efficiency than the i.p. route. The HIV-VLP(A)s represent an efficient strategy to stimulate both arms of immunity; furthermore, the induction of specific humoral immunity at mucosal sites, which nowadays represent the main port of entry for HIV-1 infection, is of great interest. All these properties, and the possible cross-clade in vivo protection, could make these HIV-VLP(A)s a good candidate for a mono- and multicomponent worldwide preventive vaccine approach not restricted to high-priority regions, such as sub-Saharan countries.     

Protection of macaques against intrarectal infection by a combination immunization regimen with recombinant simian immunodeficiency virus SIVmne gp160 vaccines

We previously reported that immunization with recombinant simian immunodeficiency virus SIVmne envelope (gp160) vaccines protected macaques against intravenous challenge by the cloned homologous virus E11S but that this protection was only partially effective against the uncloned virus, SIVmne. In the present study, we examine the protective efficacy of this immunization regimen against infection by a mucosal route. We found that the same gp160-based vaccines were highly effective against intrarectal infection not only with the E11S clone but also with the uncloned SIVmne. Protection against mucosal infection is therefore achievable by parenteral immunization with recombinant envelope vaccines. Protection appears to correlate with high levels of SIV-specific antibodies and, in animals protected against the uncloned virus, the presence of serum-neutralizing activities. To understand the basis for the differential efficacies against the uncloned virus by the intravenous versus the intrarectal routes, we examined viral sequences recovered from the peripheral blood mononuclear cells of animals early after infection by both routes. We previously showed that the majority (85%) of the uncloned SIVmne challenge stock contained V1 sequences homologous to the molecular clone from which the vaccines were made (E11S type), with the remainder (15%) containing multiple conserved changes (the variant types). In contrast to intravenously infected animals, from which either E11S-type or the variant type V1 sequences could be recovered in significant proportions, animals infected intrarectally had predominantly E11S-type sequences. Preferential transmission or amplification of the E11S-type viruses may therefore account in part for the enhanced efficacy of the recombinant gp160 vaccines against the uncloned virus challenge by the intrarectal route compared with the intravenous route.