Efficacy of HSV-1 ISCOM vaccine in the guinea-pig model of HSV-2 infection.

The capability of a herpes simplex virus (HSV)-1 ISCOM vaccine to protect against intravaginal HSV-2 challenge infection in guinea-pigs is described. The protective efficacy of the HSV-1 ISCOM vaccine is compared with that of a purified, aqueous HSV-1 antigen preparation administered using a similar immunization schedule. The results show that female guinea-pigs immunized with two doses of HSV-1 ISCOM vaccine, each consisting of 20 micrograms of protein given 2 weeks apart responded with high ELISA and neutralization antibody titres, and are almost completely protected against the clinical effects of intravaginal challenge with 10(5.2) TCID50 of HSV-2. This cross-protection is significantly greater than that observed in guinea-pigs immunized with a single dose of HSV-1 ISCOM vaccine, two doses of aqueous HSV-1 antigen preparation or two doses of a mock ISCOM vaccine. However, none of the vaccine preparations completely prevented HSV-2 replication following challenge. Western blot and radioimmunoprecipitation of sera from immunized guinea-pigs show the HSV-1 ISCOM vaccine preparation to contain the major HSV-1 glycoproteins. These findings are discussed in relation to the value and potential use of HSV-1 ISCOM vaccine in humans.     

A mixed Th1/Th2 response elicited by a liposomal formulation of Leishmania vaccine instructs Th1 responses and resistance to Leishmania donovani in susceptible BALB/c mice

In this study, we have developed a vaccine with Leishmania donovani promastigote membrane antigens (leishmanial antigens (LAg)) encapsulated in a liposome carrier formulated with distearyol (DSPC, transition temperature (Tc) = 54 degrees C) derivative of l-alpha-phosphatidyl choline, for immunizing BALB/c mice against progressive visceral leishmaniasis. This formulation could limit hepatosplenomegaly to almost normal levels and conferred strong levels of protection in both liver and spleen against challenge infection. Immunization with liposomal LAg activated peritoneal macrophages for enhanced leishmanicidal activity in association with NO production, and induced antibody as well as T-cell mediated immune responses. Production of both IFN-gamma and IL-4 by splenic T cells, and serum IgG1 and IgG2a, suggest induction of a mixed Th1/Th2 response following immunization. Experimental challenge corresponded with elevated DTH, and mitogen and antigen specific cellular responses. Increased production of NO and IFN-gamma by spleen cells, and down regulation of IL-4, demonstrate that an initial stimulation of a mixed Th1/Th2 response by vaccination instructs Th1 responses and resistance against a progressive infection by L. donovani.     

Effects of IL-12 on immune responses induced by transcutaneous immunization with antigens formulated in a novel lipid-based biphasic delivery system

The development of non-invasive methods for the delivery of proteins through the permeability barriers, such as the intact skin, will greatly facilitate the administration of human and veterinary vaccines. In the present study we used recombinant Pasteurella haemolytica leukotoxin (Lkt) and hen egg lysozyme (HEL) as model antigens to investigate the ability of transdermal administration of vaccine antigens to induce humoral and cellular responses in mice and to assess the immunomodulatory effects of IL-12 on these antigen-specific immune responses. Mice were immunized by the transdermal route with Lkt or HEL formulated in a novel lipid-based biphasic delivery system (BPDS). Transdermal delivery of Lkt or HEL induced strong polarized Th2 responses characterized by enhancement of antigen-specific IgG1 antibody subclass and predominant induction of antigen specific IL-4 over IFN-gamma in spleen and draining lymph nodes cells. Animals immunized by topical application of formulations containing antigen and IL-12 developed significantly lower antibody titres without significant changes in IL-4 or IFN-gamma secreting cells (SC) in the draining lymph nodes or spleen cells. Our results indicated that application of antigens formulated in BPDS induced antigen-specific immune responses. Furthermore, incorporation of IL-12 to the vaccine formulation influences the induction of antibody responses induced by transdermal immunization. We demonstrated the feasibility of using this technology for the development of non-invasive methods of vaccine administration.     

Interleukin-12 redirects murine immune responses to soluble or aluminum phosphate adsorbed HSV-2 glycoprotein D towards Th1 and CD4+ CTL responses

The type of immune response elicited against HSV-2 infection may be a factor in the frequency and severity of recurrent disease, with non-recurrent status being associated with a Th1-like response. As administration of glycoprotein D subunit formulated with an aluminum-based adjuvant induces predominantly Th2-like immune responses, we sought to assess the ability of IL-12 to redirect anti-HSV immunity towards a Th1 response. Co-administration of gD with IL-12 resulted in gD-specific antibody subclass switching from predominantly IgG1 observed in mice immunized with either gD or gD/AlPO4 to a more balanced combination of IgG1 and IgG2a, and enhanced virus neutralizing activity. Spleen cells from mice immunized with gD and IL-12, and restimulated in vitro with HSV-2, developed into effector cells capable of secreting IFN-gamma and lysing HSV-2 infected targets, while those obtained from gD or gD/ALPO4 immunized mice did not express lytic activity. In vitro studies determined that these CTLs were CD4+ and that the cytotoxicity was primarily perforin dependent. Vaginal challenge with HSV-2 demonstrated that IL-12 co-administration with gD resulted in increased efficacy of this vaccine as compared to administration of gD antigen alone. This acquired protection persisted up to 1 year. Finally, adsorbing gD and IL-12 to AlPO4 decreased the optimal dose of IL-12 required to enhance gD immunogenicity and shift responses towards a Th1-like profile.     

Vaccination with different HSV-1 glycoproteins induces different patterns of ocular cytokine responses following HSV-1 challenge of vaccinated mice.

We previously reported that vaccination of BALB/c mice with different baculovirus expressed HSV-1 glycoproteins induced varying degrees of protection against HSV-1 ocular challenge, ranging from complete protection to no protection, to exacerbation of eye disease. To correlate specific local immune responses with protection and exacerbation of corneal scarring, we examined immune cell infiltrates in the cornea after ocular HSV-1 challenge of vaccinated mice. Mice were vaccinated with gD, which completely protects against corneal scarring, gG, which produces no protection against corneal scarring, or gK, which exacerbates corneal scarring. Cryostat sections of cornea were taken at different times after challenge and examined for infiltrating cells containing IL-2, IL-4, IFN-gamma, IL-6, or TNF-alpha. No corneal infiltrates were seen before challenge or 1 day after ocular challenge in any groups. By days 3-7, many cells containing IL-4 and IFN-gamma, but few cells containing IL-2, had infiltrated into the corneas of gG or mock vaccinated mice. At the same times, many cells containing IL-2, but few cells containing IL-4 or IFN-gamma, were seen in the corneas of gD vaccinated mice. In contrast, the corneas of mice vaccinated with gK contained large amounts of IL-2, IFN-gamma, and IL-4. Our results suggest that: (1) corneas from gD vaccinated mice had no corneal disease and developed a response highly biased toward IL-2 responses; (2) corneas from gG or mock vaccinated eyes had significant corneal disease and developed a mostly IL-4 and IFN-gamma cytokine response; and (3) corneas from gK vaccinated mice had exacerbated corneal disease and developed strong IL-2, IL-4 and IFN-gamma cytokine responses.     

ISCOM based vaccines for cancer immunotherapy

Immunostimulating complex (ISCOM) vaccines are particulate antigen delivery vehicles composed of saponin, cholesterol, phospholipid and immunogen. Here we illustrate that ISCOM-based vaccines represent an attractive modality for the development of anti-cancer vaccines. Using murine models and a model cancer antigen, ISCOM vaccines were shown to induce potent CD8 T cell responses, to mediate protection in three different tumor models, to promote Th1-biased immunity, and to induce CD8 T cell responses in the absence of CD4+ T cell help. The former three activities were also found to be substantially improved when the vaccine antigen was associated with the ISCOM structure. Furthermore, the presence in vivo of pre-existing antibodies against the vaccine antigen did not inhibit CD8 T cell induction by the ISCOM vaccine. Although vaccination was effective against challenge with vaccine-antigen expressing tumors, no activity against neighboring vaccine-antigen negative tumor cells was observed, indicating that determinant spreading or bystander activity does not lead to significant anti-cancer activity.     

Potent protective cellular immune responses generated by a DNA vaccine encoding HSV-2 ICP27 and the E. coli heat labile enterotoxin

A mouse model was employed to evaluate protective cellular immune responses induced by an immediate early antigen of HSV-2. Particle-mediated DNA vaccination of mice with a DNA plasmid-encoding ICP27 resulted in the induction of ICP27-specific IFN-gamma and TNF-alpha production in Balb/c mice, but little protection to intranasal challenge with wild type HSV-2. However, when the DNA vaccine was supplemented with as little as 50ng of a vector encoding the A and B subunits of the Escherichia coli heat labile enterotoxin (LT), animals were profoundly protected from morbidity and mortality. The ICP27+LT-mediated protection was correlated with a large increase in ICP27-specific IFN-gamma and TNF-alpha production but cytokine-specific monoclonal antibody treatment at the time of challenge showed that protection was mediated predominantly by IFN-gamma. Furthermore, depletion of T cell subsets prior to infectious challenge demonstrated that removal of either CD8+ or CD4+ T cells impaired protection with CD8+ T cells appearing to play a direct effector role. These data demonstrate that augmented cellular immune responses resulting from LT vector plus antigen vector administration to the skin are biologically significant, leading to enhanced protection against mucosal pathogenic challenge.     

Improved T cell responses to Plasmodium falciparum circumsporozoite protein in mice and monkeys induced by a novel formulation of RTS,S vaccine antigen

Protection against Plasmodium falciparum sporozoite infection can be achieved by vaccination with the recombinant circumsporozoite protein-based vaccine RTS,S formulated with the AS02A Adjuvant System. Since this protection is only partial and wanes over time, we have developed a new RTS,S-based vaccine adjuvanted with AS01B. RTS,S/AS01B-induced high specific antibody titers and increased the frequency of mouse CD4(+) and CD8(+) T cells expressing IFN-gamma, and of monkey CD4(+) T cells expressing IL-2 and/or IFN-gamma and/or TNF-alpha upon stimulation with vaccine antigens. Our data provides clear evidence that combining RTS,S antigen with a potent adjuvant induces strong humoral and cellular responses in vivo.     

Therapeutic vaccination against HSV-2: influence of vaccine formulation on immune responses and protection in mice

Therapeutic immunisation may represent a means of influencing viral infections that persist in the host by modulating the nature or level of host immunity. To assess the influence of the form of the antigenic stimulus on immunity to type-2 herpes simplex virus (HSV-2), mice pre-infected with sublethal doses of HSV-2 were immunised with various HSV-2 vaccine formulations prior to challenge infection with heterologous HSV-1. Measurements of interleukin-2 (IL-2), interleukin-4 (IL-4) and interferon-gamma (IFN-gamma) levels in mouse spleen cell cultures restimulated in vitro with HSV-2 antigens showed that, depending on the form of HSV-2 antigen preparation used in this therapeutic context, changes in the levels of these cytokines could be effected. Measurement of HSV-specific antibody by serological tests support the contention that immunisation of HSV-2-infected mice can either enhance the existing Th1-like immune response elicited following HSV-2 infection, or modulate this response towards a more Th2-like profile, and this is dependent on the form of the antigenic stimulus. The degree of protection against subsequent lethal, heterologous HSV-1 challenge infection varied according to the nature of the infection and the immunisation history of the animals.     

Protection and serum antibody responses in guinea-pigs and mice immunized with HSV-1 antigen preparations obtained using different detergents

Guinea-pigs immunized with a zwitterionic detergent-extracted HSV-1 antigen preparation responded with EIA and NT serum antibody titres that were significantly greater than those elicited by a non-ionic detergent-extracted antigen preparation inoculated using a similar dosage schedule. Following intravaginal challenge of the guinea-pigs with HSV-2 (strain SH/B), there was no statistically significant difference in the protection afforded to these animals by the two antigen preparations, although the results indicated the zwitterionic detergent-extracted HSV-1 antigen preparation to be slightly superior in this respect. In mice, the zwitterionic detergent-extracted HSV-1 antigen preparation elicited an EIA antibody response and partial protection against homologous virus challenge. The relevance of these animal models to determination of immunogenicity and efficacy of HSV vaccines prepared for use in man, and the reasons for the differences in immunogenicity of these HSV-1 antigen preparations in guinea-pigs, are discussed.     

Inactivated HSV-2 in MPL/alum adjuvant provides nearly complete protection against genital infection and shedding following long term challenge and rechallenge

Herpes Simplex Virus Type 2 (HSV-2) infection can result in life-long recurrent genital disease, asymptomatic virus shedding, and transmission. No vaccine to date has shown significant protection clinically. Here, we used a mouse model of genital HSV-2 infection to test the efficacy of a vaccine consisting of whole, formalin-inactivated HSV-2 (FI-HSV2) formulated with monophosphoryl lipid A (MPL) and alum adjuvants. Vaccine components were administered alone or as a prime-boost immunization together with DNA vaccines encoding a truncated glycoprotein D2 (gD2t) and two conserved HSV-2 genes necessary for virus replication, UL5 (DNA helicase) and UL30 (DNA polymerase). Our results show: (1) compared with mock immunized controls, mice immunized with FI-HSV2 plus MPL/alum consistently showed protection against disease burden and total viral shedding while the mice immunized with gD2t protein with MPL/alum did not; (2) protection against genital disease and viral replication correlated with the type of boost in a prime-boost immunization with little advantage afforded by a DNA prime; (3) intramuscular (i.m.) immunization with FI-HSV2 in MPL/Alhydrogel adjuvant provided nearly complete protection against vaginal HSV-2 shedding after a lethal intravaginal (i.vag.) short-term challenge and long-term rechallenge; (4) single formulation immunization with DNA vaccines, FI-HSV2, and MPL in an aluminum phosphate (Adju-Phos) adjuvant did not increase protection relative to FI-HSV2/MPL/Adju-Phos alone; and (5) addition of MPL/alum to the FI-HSV2 was required for optimal protection against disease, viral replication, and latent virus load in the dorsal root ganglia (DRG). Most notably, an optimized vaccine formulation of FI-HSV2 MPL/Alhydrogel given i.m. completely protected against detectable vaginal HSV-2 shedding in the majority of animals and HSV-2 latent DNA in the DRG of all animals.     

DNA vaccination with map1 gene followed by protein boost augments protection against challenge with Cowdria ruminantium, the agent of heartwater.

A DNA vaccine encoding the immunodominant MAP1 protein of Cowdria ruminantium (Crystal Springs (CS) strain) was shown to partially protect DBA/2 mice against homologous lethal challenge. To enhance the protective capacity of this DNA vaccine, the effects of length of interval between vaccinations and of prime-boost regimes were investigated. Increasing the interval between vaccinations from 2 to 12 weeks did not result in better protection (P=0.900). However, boosting DNA vaccine-primed mice with recombinant MAP1 protein significantly augmented protection on homologous challenge in various trials from 13-27 to 53-67% (P<0.050). The augmented protection by the prime-boost regimen correlated with augmented T(H1) type immune responses that were induced by the DNA vaccine. These responses were characterized by production of IFN-gamma, IL-2 and anti-MAP1 antibodies of predominantly IgG2a isotype, and were critical for protection against C. ruminantium infection. Cytokine analyses were done at 48h after in vitro stimulation of splenocytes with C. ruminantium or control antigens. In contrast, splenocytes of DNA vector control mice produced no cytokines and these mice were fully susceptible to challenge. In addition, DBA/2 mice immunized with the recombinant MAP1 protein without DNA vaccine priming produced non-protective T(H2) type immune responses which were characterized by production of IL-4, IL-5, IL-10 and IgG1 anti-MAP1 antibodies. A second DNA vaccine containing map1 gene from the Mbizi strain of C. ruminantium also delivered by a prime-boost regime, conferred less protection against heterologous challenge. Hence, in developing DNA vaccines against heartwater that contain map1 gene, a prime-boost regimen should be adopted and gene sequence heterogeneity of field isolates should also be considered.     

Vaccination of mice with a cocktail DNA vaccine induces a Th1-type immune response and partial protection against Schistosoma japonicum infection.

Several defined vaccine candidate antigens of Schistosoma japonicum have shown promise in large animal vaccination experiments. However, vaccination of mice in the laboratory with either single recombinant antigens or DNA encoding forms of the individual antigens has so far failed to induce significant protection against S. japonicum cercarial challenge infection as judged by worm reduction, although specific antibodies were generated. This is in contrast to the results achieved using radiation-attenuated vaccines which are highly protective. Even in large animal vaccination experiments, the protection levels obtained with single defined antigens were far below those achieved using the attenuated vaccines. One possible interpretation is that the immune responses induced by single antigen vaccination may not be strong enough to combat the challenging infection. We, therefore, carried out mouse vaccination experiments using a cocktail DNA vaccine comprising four DNA plasmids encoding four different S. japonicum antigens, Sj62, Sj28, Sj23 and Sj14-3-3, respectively. We, also investigated whether co-injection of the mouse IL-12 encoding plasmid with the cocktail DNA vaccine was able to enhance the Th1 responses and hence the protective immunity. Three intramuscular injections of the cocktail DNA vaccine induced a significant Th1-type cellular response with high level of IFN-gamma production by splenocytes upon in vitro stimulation with recombinant antigens. Importantly, significant IgG antibody responses were also induced against crude worm antigens. In two out of three experiments, significant resistance (34-37 and 44-45%, respectively) was demonstrated while another experiment did not show any protection against S. japonicum cercarial challenge infection. Co-injection of the IL-12 encoding DNA did not further enhance these responses, nor the level of resistance, compared with the cocktail DNA alone.     

Biochemical characterization of herpes simplex virus type-1-immunostimulating complexes (ISCMOs): a multi-glycoprotein structure.

The preparation and characterization of an immunostimulating complex (ISCOM) preparation containing several HSV-1 glycoproteins, including the major glycoproteins B and D is described. The multi-glycoprotein HSV-1 ISCOM preparation was obtained from a gradient-purified aqueous HSV-1 antigen preparation following extraction from infected cells using a zwitterionic detergent. With polyclonal and monoclonal antibodies to HSV-1 glycoproteins in enzyme-linked immunosorbent assay, SDS-polyacrylamide gel electrophoresis and radioimmunoprecipitation techniques, the HSV-1 ISCOM preparation was shown to contain glycoproteins B, C, D, E, H and I, although further, additional proteins were also present. The DNA content of HSV-1 ISCOMs was determined using a 3H-thymidine labelling method. The protein and DNA contents of the HSV-1 ISCOM preparation are discussed with reference to the potentialities of the preparation as a vaccine for use in human beings.     

Co-administration of certain DNA vaccine combinations expressing different H5N1 influenza virus antigens can be beneficial or detrimental to immune protection

Achieving broad-spectrum immunity against emerging zoonotic viruses such as avian influenza H5N1 and other possible pandemic viruses will require generation of cross-protective immune responses. Strong antibody responses generated against the H5HA protein are protective, however, antigenic variation between diverging isolates can interfere with virus neutralization. The current study investigates co-administration of an H5 HA DNA vaccine with other variable and conserved influenza antigens (NA, NP, and M2). All antigens were derived from the A/Hanoi/30408/2005 (H5N1) virus and the contribution towards overall protection and immune activation was assessed against lethal homologous and heterologous challenges. An (HA + NA) combination afforded the best protection against homologous challenge and (HA + NP) was comparable to HA alone against heterologous A/Hong Kong/483/1997 challenge. Interestingly, combining all four H5 antigens at a single site did not improve protection against matched challenge and unexpectedly reduced survival by 30% against a heterologous challenge. Survival was also significantly decreased against heterologous challenge following combination of (HA + NP) with an unrelated antigen. Although there were no significant changes in antibody titres, significantly lower T-cell responses were detected against all antigens except HA in each combination. Co-administration of the vaccines at different injection sites restored T-cell responses but did not improve overall protection. Similar observations were also recorded following combination of HA and NP antigens using two different adenovirus-based backbones. Overall, the data suggest that co-administering certain H5N1 antigens offer better or comparable protection to HA alone, however, combining extra antigens may be unnecessary and lead to unfavourable immune responses.     

Protective Th1 immune responses against chronic toxoplasmosis induced by a protein-protein vaccine combination but not by its DNA-protein counterpart

Vaccine-induced protection against toxoplasmosis is correlated with cellular immune responses to Toxoplasma gondii, both in animals and man. The goal of the current study was to evaluate whether the combination of a recombinant protein and a plasmid DNA vaccine could offer an advantage over the protein mixture, and protect outbred mice against infection with T. gondii. To this purpose, the chimeric protein rEC2, encoding antigenic fragments of surface-associated proteins MIC2, MIC3 and SAG1, was combined with pGRA7 plasmid DNA or rGRA7 protein. High levels of antibodies were elicited by both vaccine formulations. The protein-DNA vaccine elicited a polarized Th1/Th2 immune response, characterized by IFN-gamma and IL-10, and afforded low protection (24%) against brain cyst formation. In contrast, the protein-protein vaccine elicited a Th1-focused immune response, characterized by IFN-gamma and IL-2 production, conferring a strong protection (79%) against brain cyst formation in chronic toxoplasmosis. We show here that GERBU adjuvanted protein vaccines confer better protection against toxoplasmosis than the protein-DNA heterologous vaccine.     

Oral immunisation with live aroA attenuated Salmonella enterica serovar Typhimurium expressing the Yersinia pestis V antigen protects mice against plague

Bubonic and pneumonic plague are caused by the bacterium Yersinia pestis. The V antigen of Y. pestis is a protective antigen against plague. In this study, an aroA attenuated strain of Salmonella enterica serovar Typhimurium (SL3261) has been used to deliver the Y. pestis V antigen as a candidate oral plague vaccine. SL3261 was transformed with the expression plasmid pTrc-LcrV, containing the lcrV gene encoding V antigen. Immunoblot analysis showed V antigen expression in SL3261 in vitro and intragastric immunisation of mice with the recombinant Salmonella resulted in the induction of V antigen-specific serum antibody responses and afforded protection against Y. pestis challenge. However, the antibody responses induced by the recombinant Salmonella did not correlate with the protection afforded, indicating that immune responses other than antibody may play a role in the protection afforded against plague by this candidate vaccine.     

Prolonged expression of IFNgamma induced by protective blood-stage immunization against Plasmodium yoelii malaria.

Mice vaccinated with whole blood-stage antigens of Plasmodium yoelii develop protective, antibody-mediated immune responses to homologous challenge infection. In this model the level of protection induced by whole parasite antigen vaccination is dependent on antibody isotype, which can be influenced by adjuvant formulations. In this study the ability adjuvant formulations to affect cytokine production and protection against P. yoelii blood-stage infection was investigated. Survival of mice in groups vaccinated with P. yoelii antigens in an aqueous mix of copolymer P1005 + RaLPS was 100%. Mice vaccinated with either P. yoelii antigens alone or combined with a water-in-oil emulsion of copolymer P1005 + RaLPS demonstrated 83 or 50% survival, respectively. The fully protective aqueous vaccine group produced higher levels of interferon gamma (IFNgamma) and interleukin 4 (IL-4) than the water-in-oil vaccine group following a live parasite challenge infection. Furthermore, mice vaccinated with the aqueous vaccine displayed prolonged IFNgamma and IL-4 response as compared to mice that received the same antigens without adjuvants. These data support the hypothesis that both the Th1 cytokine IFNgamma, and the Th2 cytokine IL-4 are modulated by the vaccine vehicle and adjuvant used for vaccination, thus possibly affecting expression of protective immune responses. However, it is the long-lasting IFNgamma response following blood-stage P. yoelii parasite challenge that is associated with enhanced survival.     

In vivo activity of cationic immune stimulating complexes (PLUSCOMs)

A particulate vaccine delivery system consisting of cationic ISCOM derivatives (PLUSCOMs) was compared to classic anionic ISCOMs with regard to antigen attachment and ability to elicit in vivo T cell responses against a model protein antigen (ovalbumin [OVA]). ISCOMs did not incorporate hydrophilic OVA whilst OVA readily adsorbed onto PLUSCOMs with increasing adsorption at higher protein concentrations. The zeta-potential of PLUSCOMs significantly decreased with increasing protein load, suggesting neutralization of the cationic charge upon absorption of the anionic OVA. Antigen-specific CD8 T cell responses were demonstrated in mice vaccinated with either PLUSCOMs or ISCOMs. Ex vivo restimulation of harvested T cells demonstrated that cells isolated from PLUSCOM and ISCOM vaccinated mice responded to the secondary OVA challenge more efficiently than mice vaccinated with OVA in solution. Restimulated cells from the mice vaccinated with particulate vaccines produced significantly more INF-gamma. Therefore PLUSCOMs are as effective as classic ISCOMs in inducing antigen-specific CD8 T cell responses and have advantages with regard to the incorporation of purified anionic antigens.     

Interleukin-12 redirects murine immune responses to soluble or aluminum phosphate adsorbed HSV-2 glycoprotein D towards Th1 and CD4+ CTL responses

The type of immune response elicited against HSV-2 infection may be a factor in the frequency and severity of recurrent disease, with non-recurrent status being associated with a Th1-like response. As administration of glycoprotein D subunit formulated with an aluminum-based adjuvant induces predominantly Th2-like immune responses, we sought to assess the ability of IL-12 to redirect anti-HSV immunity towards a Th1 response. Co-administration of gD with IL-12 resulted in gD-specific antibody subclass switching from predominantly IgG₁ observed in mice immunized with either gD or gD/AlPO₄ to a more balanced combination of IgG₁ and IgG_2a, and enhanced virus neutralizing activity. Spleen cells from mice immunized with gD and IL-12, and restimulated in vitro with HSV-2, developed into effector cells capable of secreting IFN-γ and lysing HSV-2 infected targets, while those obtained from gD or gD/ALPO₄ immunized mice did not express lytic activity. In vitro studies determined that these CTLs were CD4⁺ and that the cytotoxicity was primarily perforin dependent. Vaginal challenge with HSV-2 demonstrated that IL-12 co-administration with gD resulted in increased efficacy of this vaccine as compared to administration of gD antigen alone. This acquired protection persisted up to 1 year. Finally, adsorbing gD and IL-12 to AlPO₄ decreased the optimal dose of IL-12 required to enhance gD immunogenicity and shift responses towards a Th1-like profile.