HSV-2 Challenge Research Articles

Impact of CD4+ T lymphocytes on the cellular and molecular milieu of the vaginal mucosa following HSV-2 challenge of immune guinea pigs

CD4+ and CD8+ tissue resident memory cells (TRM) express many shared anti-viral activities upon re-exposure to virus. CD4+ T cells were depleted from HSV-immune guinea pigs to identify CD4-dependent functions in the vaginal mucosa following HSV-2 challenge. The incidence of animals shedding HSV-2 fell rapidly after challenge in control animals but remained significantly higher through day four post infection in CD4-depleted animals. Genes encoding CD14, IFN-γ, CCL2, and CCL5 were up-regulated in the vaginal mucosa of both groups following challenge. However, significantly higher expression of CD107b, IL-15, and TLR9 but lower expression of CD20, IL-21, and CCL5 was detected in CD4-depleted- compared to control-treated animals. Further, antigen stimulation of CD4+ TRM increased the expression of IFN-γ, IL-2, IL-21, IL-17A, and CCL5. The impact of these gene expression patterns on the recruitment and maintenance of the cellular milieu of the vaginal mucosa upon virus challenge is discussed.

Investigating the Distinct Tissue Repair and Immunosuppressive Roles of Regulatory T cells in HSV-2 Infection

Abstract Herpes Simplex Virus 2 (HSV-2) is the primary cause of genital herpes, affecting 500 million people worldwide of which approximately 64% are women. Despite this disproportionately high burden, the female reproductive tract and its unique mucosal immune environment is surprisingly understudied. Tregs are necessary for CD4 +T cell priming in the draining lymph node (dLN), though their role in shaping tissue-resident memory T cell (T rm) responses during HSV-2 reactivation is unclear. Our preliminary data indicate that activated Tregs accumulate and persist in mouse vaginal tissue (VT) after above baseline out to at least 90 days post-infection (p.i.), thus implicating Tregs in memory recall responses to reinfection. We hypothesize that Tregs restrain robust T rmrecall responses to HSV-2 challenge. Additionally, we hypothesize that vaginal Tregs promote tissue healing in the VT in response to inflammatory alarmins through the expression of the epidermal growth factor ligand, amphiregulin (Areg). Female FoxP3 DTRmice were infected intravaginally with attenuated HSV-2, then systemically depleted of Tregs via intraperitoneal injection of diphtheria toxin and challenged with WT HSV-2 on day 30 p.i. Systemically Treg-depleted mice show increased activation and proliferation of T cells, and increased frequency of cytotoxic HSV-2-specific CD8 +T cells in the VT by day 3 post-challenge. Treg-depleted mice also have significantly higher pathology scores in H&E-stained VT. We also report that VT Tregs from infected mice express more Areg than dLN Tregs via flow cytometry and single cell RNAseq. Ongoing work includes characterization of Areg expressing VT Tregs and further dissection of the T rmrecall responses in Treg-depleted mice. NIH (T32 AI07140, R01 AI141435)

Primary oral vaccination followed by a vaginal pull protects mice against genital HSV-2 infection.

HSV-2 infected more than 491 million people aged 15-49 world-wide in 2016. The morbidity associated with recurrent infections and the increased risk of HIV infection make this a major health problem. To date there is no effective vaccine. Because HSV-2 ascends to the dorsal route ganglion within 12-18 hours of infection, an effective vaccine will need to elicit a strong local resident CD8+ T cell response to prevent the infection from becoming life-long. Using a mouse model we investigated the potential of oral immunization with a novel lipid adjuvant (LiporaleTM) followed by local vaginal application of an inflammatory agents to protect against primary HSV-2 infections. Oral vaccination of mice with live-attenuated HSV-2 in LiporaleTM followed by vaginal application of DNFB or CXCL9/10 led to recruitment of tissue-resident CD8+ memory cells into the genital epithelia. This prime and pull vaccination strategy provided complete protection against wild-type HSV-2 challenge and prevented viral dissemination to the spinal cords. Activation of mucosal immunity by oral immunization, combined with induction of transient local genital inflammation can recruit long-lived tissue resident CD8+ T cells into the genital epithelium, providing significant protection against primary HSV-2 infection. This article is protected by copyright. All rights reserved.

CCL28 Enhances HSV-2 gB-Specific Th1-Polarized Immune Responses against Lethal Vaginal Challenge in Mice.

Plasmid DNA (pDNA) represents a promising “genetic vaccine platform” capable of overcoming major histocompatibility complex barriers. We previously demonstrated that low-to-moderate doses of mucosae-associated epithelial chemokine (MEC or CCL28) as an immunomodulatory adjuvant can trigger effective and long-lasting systemic and mucosal HSV-2 gD-specific immune responses, whereas mice immunized with gD in combination with high-dose CCL28 showed toxicity and lost their immunoprotective effects after lethal HSV-2 challenge. The exact causes underlying high-dose, CCL28-induced lesions remain unknown. In an intramuscularly immunized mouse model, we investigated the immune-enhancement mechanisms of low-dose CCL28 as a molecular adjuvant combined with the relatively weak immunogen HSV-2 gB. Compared with the plasmid gB antigen group, we found that a low-dose of plasmid CCL28 (pCCL28) codelivered with pgB induced increased levels of gB-specific serum IgG and vaginal fluid IgA, serum neutralizing antibodies (NAb), Th1-polarized IgG2a, and cytokine IL-2 (>5-fold). Furthermore, low-dose pCCL28 codelivery with pgB enhanced CCL28/CCR10-axis responsive CCR10− plus CCR10+ B-cell (~1.2-fold) and DC pools (~4-fold) in the spleen, CCR10− plus CCR10+ T-cell pools (~2-fold) in mesenteric lymph nodes (MLNs), and the levels of IgA-ASCs in colorectal mucosal tissues, leading to an improved protective effect against a lethal dose of HSV-2 challenge. Findings in this study provide a basis for the development of CCL28-adjuvant vaccines against viral mucosal infections.

Bystander CD8 T cell memory responses partially protect mice against lethal vaginal HSV-2 challenge

Abstract Herpes Simplex Virus type-2 (HSV-2) infection is one of the most prevalent sexually transmitted infections, yet no vaccine is currently available. Detailed analysis of HSV-infected human tissue revealed tissue-resident CD8 T cells (TRM) limit the duration and severity of HSV-2 episodes. Interestingly, recent studies indicate that the sensing and alarming function of CD8 TRMs is not restricted to cognate antigen interaction, but CD8 TRM can mediate protection against antigenically unrelated pathogens, termed “bystander activation.” Here, we investigated if antigen non-specific CD8 T cells could provide some degree of protection in a bystander fashion in the context of HSV infection. To test this, we created antigen non-specific memory compartments through immunization of mice with Listeria expressing ovalbumin (OVA) (LM-OVA). Mice were then challenged with wild-type HSV-2 to assess the degree of bystander-mediated protection. Immunization with LM-OVA-delayed disease progression from lethal viral challenge, suggesting that bystander CD8 T cells may mediate protection despite the lack of antigen-specificity. Furthermore, we found early infiltration of antigen-non-specific CD8 T cells (OVA-specific) in the vaginal tract in the infection setting. Additionally, assessment of the memory CD8 compartment in human vaginal tissue upon in-vitro treatment with cytokines caused bystander activation of memory CD8 T cells. Finally, we found that in-vitro treatment of CD8 T cells with cytokines delayed disease progression and reduced viral burden upon adoptive transfer. Altogether, our findings suggest that local bystander-activation of CD8 memory T cells can participate in protection from HSV-2.

Bystander CD8 T cell memory responses partially protect mice against lethal vaginal HSV-2 challenge

Abstract Herpes Simplex Virus type-2 (HSV-2) infection is one of the most prevalent sexually transmitted diseases, yet no vaccine is currently available. Detailed analysis of HSV-infected human tissue revealed tissue-resident CD8 T cells (TRM) limit the duration and severity of HSV-2 episodes. Interestingly, recent studies reveal that the sensing and alarming function of CD8 TRMs is not restricted to cognate antigen interaction, but CD8 TRM can mediate protection against antigenically unrelated pathogens, termed “bystander activation.” Here, we extended our findings to determine if antigen non-specific CD8 T cells could provide some degree of protection in a bystander fashion in the context of HSV infection. To test this, we created antigen non-specific memory compartments through immunization of mice with Listeria expressing ovalbumin (OVA) (LM-OVA). Mice were then challenged with wild-type HSV-2 to assess the degree of vaccine-mediated protection. Immunization with LM- OVA-induced intermediate protection from lethal viral challenge, suggesting that bystander CD8 T cells (BA-CTL) may mediate protection despite the absence of antigen-specificity. The bystander functionality could be context-dependent, and the unregulated expansion of these cells can be damaging to the host. A potential explanation of BA-CTL regulation is via the involvement of regulatory T cells (Tregs), in the absence of Treg-mediated regulation, massive expansion of BA-CTLs can be predicted. On the contrary, we found depletion of Tregs resulted in significantly fewer BA- CTLs in the vaginal tract on days 2 and 6 post-HSV-2 challenge. Hence how these cells maintain the delicate balance of protection against pathology necessitates further investigation.

An HSV-2 nucleoside-modified mRNA genital herpes vaccine containing glycoproteins gC, gD, and gE protects mice against HSV-1 genital lesions and latent infection.

HSV-1 causes 50% of first-time genital herpes infections in resource-rich countries and affects 190 million people worldwide. A prophylactic herpes vaccine is needed to protect against genital infections by both HSV-1 and HSV-2. Previously our laboratory developed a trivalent vaccine that targets glycoproteins C, D, and E present on the HSV-2 virion. We reported that this vaccine protects animals from genital disease and recurrent virus shedding following lethal HSV-2 challenge. Importantly the vaccine also generates cross-reactive antibodies that neutralize HSV-1, suggesting it may provide protection against HSV-1 infection. Here we compared the efficacy of this vaccine delivered as protein or nucleoside-modified mRNA immunogens against vaginal HSV-1 infection in mice. Both the protein and mRNA vaccines protected mice from HSV-1 disease; however, the mRNA vaccine provided better protection as measured by lower vaginal virus titers post-infection. In a second experiment, we compared protection provided by the mRNA vaccine against intravaginal challenge with HSV-1 or HSV-2. Vaccinated mice were totally protected against death, genital disease and infection of dorsal root ganglia caused by both viruses, but somewhat better protected against vaginal titers after HSV-2 infection. Overall, in the two experiments, the mRNA vaccine prevented death and genital disease in 54/54 (100%) mice infected with HSV-1 and 20/20 (100%) with HSV-2, and prevented HSV DNA from reaching the dorsal root ganglia, the site of virus latency, in 29/30 (97%) mice infected with HSV-1 and 10/10 (100%) with HSV-2. We consider the HSV-2 trivalent mRNA vaccine to be a promising candidate for clinical trials for prevention of both HSV-1 and HSV-2 genital herpes.

IL-36γ Is a Key Regulator of Neutrophil Infiltration in the Vaginal Microenvironment and Limits Neuroinvasion in Genital HSV-2 Infection.

HSV-2 is a neurotropic virus that causes a persistent, lifelong infection that increases risk for other sexually transmitted infections. The vaginal epithelium is the first line of defense against HSV-2 and coordinates the immune response through the secretion of immune mediators, including the proinflammatory cytokine IL-36γ. Previously, we showed that IL-36γ treatment promoted transient polymorphonuclear cell infiltration to the vaginal cavity and protected against lethal HSV-2 challenge. In this report, we reveal that IL-36γ specifically induces transient neutrophil infiltration but does not impact monocyte and macrophage recruitment. Using IL-36γ-/- mice in a lethal HSV-2 challenge model, we show that neutrophil counts are significantly reduced at 1 and 2 d postinfection and that KC-mediated mature neutrophil recruitment is impaired in IL-36γ-/- mice. Additionally, IL-36γ-/- mice develop genital disease more rapidly, have significantly reduced survival time, and exhibit an increased incidence of hind limb paralysis that is linked to productive HSV-2 infection in the brain stem. IL-36γ-/- mice also exhibit a significant delay in clearance of the virus from the vaginal epithelium and a more rapid spread of HSV-2 to the spinal cord, bladder, and colon. We further show that the decreased survival time and increased virus spread observed in IL-36γ-/- mice are not neutrophil-dependent, suggesting that IL-36γ may function to limit HSV-2 spread in the nervous system. Ultimately, we demonstrate that IL-36γ is a key regulator of neutrophil recruitment in the vaginal microenvironment and may function to limit HSV-2 neuroinvasion.

The Murine Intravaginal HSV-2 Challenge Model for Investigation of DNA Vaccines.

DNA vaccines have been licensed in veterinary medicine and have promise for humans. This format is relatively immunogenic in mice and guinea pigs, the two principle HSV-2 animal models, permitting rapid assessment of vectors, antigens, adjuvants, and delivery systems. Limitations include the relatively poor immunogenicity of naked DNA in humans and the profound differences in HSV-2 pathogenesis between host species. Herein, we detail lessons learned investigating candidate DNA vaccines in the progesterone-primed female mouse vaginal model of HSV-2 infection as a guide to investigators in the field.

IL-36γ regulates neutrophil infiltration and limits neuroinvasion in genital HSV-2 pathogenesis

Abstract Herpes simplex virus 2 is a neurotropic virus that causes a persistent, life-long infection that increases risk for other sexually transmitted infections. The vaginal epithelium is the first line of defense against HSV-2 and secretes immune mediators, including the pro-inflammatory cytokine IL-36γ, to coordinate the immune response. Previously, we showed that IL-36γ treatment promoted polymorphonuclear cell infiltration to the vaginal cavity and protected against lethal HSV-2 challenge. Herein, we extended these findings using flow cytometry to profile the cellular infiltrate. IL-36γ significantly (p < 0.0001) induced transient neutrophil infiltration, but did not impact monocyte or macrophage recruitment. Using IL-36γ−/− mice in a lethal HSV-2 challenge model, we show that neutrophils were significantly (p < 0.0001) depleted at 1- and 2-days post infection, and that mature neutrophil recruitment was impaired. In addition, IL-36γ−/− mice develop genital disease more rapidly (p < 0.0001), have significantly (p < 0.0001) reduced survival time, and display an increased incidence (p < 0.0001) of neurologic disease compared to wild type mice. IL-36γ−/− mice exhibited a significant (p < 0.05) delay in clearance of the virus from the vaginal epithelium, and analysis of virus spread to the spinal cord, bladder, and colon demonstrated that HSV-2 spread significantly (p < 0.05) more quickly in IL-36γ−/− mice. Further, we detected productive HSV-2 infection in the brainstem of IL-36γ−/− mice significantly (p < 0.0001) more frequently than in wild type mice. Ultimately, we demonstrate that IL-36γ is a key regulator of neutrophil recruitment in the vaginal microenvironment and may function to limit HSV-2 neuroinvasion.

The type I interferon receptor is not required for protection in the Chlamydia muridarum and HSV-2 murine super-infection model.

Chlamydia trachomatis/HSV-2 vaginal co-infections are seen clinically, suggesting that these sexually transmitted pathogens may interact. We previously established an intravaginal Chlamydia muridarum/HSV-2 super-infection model and observed that chlamydial pre-infection protects mice from a subsequent lethal HSV-2 challenge. However, the mechanism of protection remains unknown. The type I interferon, IFN-β, binds to the type I interferon receptor (IFNR), elicits a host cellular antiviral response and inhibits HSV replication in vitro and in vivo. Previous studies have demonstrated that C. muridarum infection stimulates genital tract (GT) IFN-β production; therefore, we hypothesized that chlamydial pre-infection protects mice from HSV-2 challenge via the IFN-β/IFNR-induced antiviral response. To test this prediction, we quantified IFN-β levels in vaginal swab samples. Detection of IFN-β in C. muridarum singly infected, but not in mock-infected animals, prompted the use of the super-infection model in IFNR knockout (IFNR−/−) mice. We observed that C. muridarum pre-infection reduces HSV-2-induced mortality by 40% in wild-type mice and by 60% IFNR−/− mice. Severity of HSV-2 disease symptoms and viral shedding was also similarly reduced by C. muridarum pre-infection. These data indicate that, while chlamydial infection induces GT production of IFN-β, type I IFN-induced antiviral responses are likely not required for the observed protective effect.

Intradermal immunisation using the TLR3-ligand Poly (I:C) as adjuvant induces mucosal antibody responses and protects against genital HSV-2 infection.

Development of vaccines able to induce mucosal immunity in the genital and gastrointestinal tracts is a major challenge to counter sexually transmitted pathogens such as HIV-1 and HSV-2. Herein, we showed that intradermal (ID) immunisation with sub-unit vaccine antigens (i.e., HIV-1 gp140 and HSV-2 gD) delivered with Poly(I:C) or CpG1668 as adjuvant induces long-lasting virus-specific immunoglobulin (Ig)-G and IgA antibodies in the vagina and feces. Poly(I:C)-supplemented sub-unit viral vaccines caused minimal skin reactogenicity at variance to those containing CpG1668, promoted a delayed-type hypersensitivity (DTH) to the vaccine and protected mice from genital and neurological symptoms after a lethal vaginal HSV-2 challenge. Interestingly, Poly(I:C12U) (Ampligen), a Poly(I:C) structural analogue that binds to TLR3 but not MDA-5, promoted robust mucosal and systemic IgG antibodies, a weak skin DTH to the vaccine but not IgA responses and failed to confer protection against HSV-2 infection. Moreover, Poly(I:C) was far superior to Poly(I:C12U) at inducing prompt and robust upregulation of IFNß transcripts in lymph nodes draining the injection site. These data illustrate that ID vaccination with glycoproteins and Poly(I:C) as adjuvant promotes long-lasting mucosal immunity and protection from genital HSV-2 infection, with an acceptable skin reactogenicity profile. The ID route thus appears to be an unexpected inductive site for mucosal immunity and anti-viral protection suitable for sub-unit vaccines. This works further highlights that TLR3/MDA5 agonists such as Poly(I:C) may be valuable adjuvants for ID vaccination against sexually transmitted diseases.

Estradiol Enhances CD4+ T-Cell Anti-Viral Immunity by Priming Vaginal DCs to Induce Th17 Responses via an IL-1-Dependent Pathway.

Clinical and experimental studies have shown that estradiol (E2) confers protection against HIV and other sexually transmitted infections. Here, we investigated the underlying mechanism. Better protection in E2-treated mice, immunized against genital HSV-2, coincided with earlier recruitment and higher proportions of Th1 and Th17 effector cells in the vagina post-challenge, compared to placebo-treated controls. Vaginal APCs isolated from E2-treated mice induced 10-fold higher Th17 and Th1 responses, compared to APCs from progesterone-treated, placebo-treated, and estradiol-receptor knockout mice in APC-T cell co-cultures. CD11c+ DCs in the vagina were the predominant APC population responsible for priming these Th17 responses, and a potent source of IL-6 and IL-1β, important factors for Th17 differentiation. Th17 responses were abrogated in APC-T cell co-cultures containing IL-1β KO, but not IL-6 KO vaginal DCs, showing that IL-1β is a critical factor for Th17 induction in the genital tract. E2 treatment in vivo directly induced high expression of IL-1β in vaginal DCs, and addition of IL-1β restored Th17 induction by IL-1β KO APCs in co-cultures. Finally, we examined the role of IL-17 in anti-HSV-2 memory T cell responses. IL-17 KO mice were more susceptible to intravaginal HSV-2 challenge, compared to WT controls, and vaginal DCs from these mice were defective at priming efficient Th1 responses in vitro, indicating that IL-17 is important for the generation of efficient anti-viral memory responses. We conclude that the genital mucosa has a unique microenvironment whereby E2 enhances CD4+ T cell anti-viral immunity by priming vaginal DCs to induce Th17 responses through an IL-1-dependent pathway.

Herpes simplex virus-2 ΔgD vaccination primes robust cellular immunity with memory Th17 cell recall following challenge

Abstract Approximately 25 percent of women are infected with herpes simplex virus (HSV)-2 in the United States. Even with anti-viral treatments, the rate of HSV transmission has increased 30 percent in the last 20 years. Furthermore the risk of acquiring an HIV infection increases 2–4 fold for HSV-2-infected individuals. Therefore, the need to have an FDA-approved HSV vaccine is immediate. Recently we have developed a single-cycle HSV-2 vaccine strain genetically deficient in glycoprotein D (HSV-2 ΔgD). Complete protection from disease and the establishment of latency is observed following either HSV-1 or HSV-2 challenge of HSV-ΔgD-vaccinated mice. We have also previously demonstrated that HSV-2 ΔgD elicits a protective, broadly functional antibody response. We hypothesized that HSV-2 ΔgD vaccination also primed T cell immunity, contributing to vaccine-induced protection. We observe both CD4 and CD8 T cells are activated by vaccination, and contract to establish a stable memory population. Vaccine-induced memory CD8 T cells are polyfunctional cytokine-producers with 55 percent specific for the immunodominant HSV gB498-505 epitope. Memory CD4 T cells primed by vaccination are quickly recalled to the infection site following challenge and exhibit a Th17-dominant phenotype with 60 percent of recalled cells producing IL-17A. We also determined that CD4 T cell help is required to stabilize the germinal center B cell population and to elicit protective antibodies following vaccination. Together, our data demonstrate HSV-2 ΔgD vaccination induces robust cellular, as well as humoral, immunity and likely an ideal vaccine vector.

Protection from genital herpes disease, seroconversion and latent infection in a non-lethal murine genital infection model by immunization with an HSV-2 replication-defective mutant virus

Viral vaccines have traditionally protected against disease, but for viruses that establish latent infection, it is desirable for the vaccine to reduce infection to reduce latent infection and reactivation. While seroconversion has been used in clinical trials of herpes simplex virus (HSV) vaccines to measure protection from infection, this has not been modeled in animal infection systems. To measure the ability of a genital herpes vaccine candidate to protect against various aspects of infection, we established a non-lethal murine model of genital HSV-2 infection, an ELISA assay to measure antibodies specific for infected cell protein 8 (ICP8), and a very sensitive qPCR assay. Using these assays, we observed that immunization with HSV-2 dl5-29 virus reduced disease, viral shedding, seroconversion, and latent infection by the HSV-2 challenge virus. Therefore, it may be feasible to obtain protection against genital disease, seroconversion and latent infection by immunization, even if sterilizing immunity is not achieved.

52. Gorilla Adenovirus Vectors for Molecular Therapeutics and Vaccines

Adenovirus vectors built from gorilla adenovirus are showing superior performance characteristics for molecular vaccines. These new adenovectors (GC Ads) are built from virus isolated from gorilla, and have been cloned and engineered to be replication-defective. GC Ads grow efficiently to high titer on cell lines GenVec has characterized, banked and filed with regulatory agencies for scalable manufacturing. They can be deleted of multiple regions (e.g., E1, E3, and E4) to provide space for multiple transgene cassettes, provide multiple blocks to eliminate replication competent virus and to ensure safer transgene delivery. These new adenovectors have been shown to not be recognized by human sera from either the United States or from Africa. A single intramuscular injection of these GC Ads can induce durable immune responses to encoded antigen. For example, intramuscular injection of GC Ads expressing RSV fusion (F0) glycoprotein induced neutralizing antibody titers that were durable > 24weeks, and protected the lung and nose against RSV challenge at even low doses of immunization in cotton rats. A single intramuscular administration of GC Ads expressing the malaria antigen CSP generated CD8 T cell and antibody responses to CSP that were greater than those generated with Ad5 in mice. In a mouse malaria challenge model the GC Ad regimen was more protective than Ad5. A single intramuscular administration of GC Ads expressing two HSV antigens, UL19 and UL47, resulted in high level cellular responses that were protective against intravaginal HSV2 challenge in mice and the CD8 T cell responses were shown to be durable, 26 weeks. Moreover, repeat administration of the same GC Ad resulted in a boost of antigen-specific immune response. Unlike other adenoviral vectors, homologous prime boost regimens with these GC Ads resulted in a boost of the CD8 T cell responses at all doses and intervals tested. These new GC Ads based on gorilla adenoviruses show generation of strong, durable T cell and neutralizing antibody responses to multiple antigens from a single administration that can be further enhanced by repeat administration. Based on these data, the platform shows great promise for development of a broad array of immunotherapeutic and molecular vaccine product candidates.

Single dose of Glycoprotein K (gK)-deleted HSV-1 live-attenuated virus protects mice against lethal vaginal challenge with HSV-1 and HSV-2 and induces lasting T cell memory immune responses

BackgroundHerpes simplex virus type-1(HSV-1) and HSV-2 are important human pathogens that cause significant ocular and urogenital complications, respectively. We have previously shown that HSV-1 virions lacking glycoprotein K (gK) are unable to enter into neurons via synaptic axonal membranes and be transported in either retrograde or anterograde manner. Here, we tested the ability of HSV-1 (F) gK-null to protect against lethal challenge with either highly virulent ocular HSV-1 (McKrae strain), or genital HSV-2 (G strain). The gK-null virus vaccine efficiently protected mice against lethal vaginal infection with either HSV-1(McKrae) or HSV-2 (G).ResultsFemale mice were immunized via a single intramuscular injection with 106 PFU of the gK-null virus. Immunized mice were treated with Depo-Provera fourteen days after vaccination and were challenged via the vaginal route one week later. Ninety percent of mice vaccinated with the gK-null virus survived HSV-1 (McKrae) challenge, while 70% of these mice survived after HSV-2 (G) challenge. Moreover, all vaccinated mice exhibited substantially reduced disease symptoms irrespective of HSV-1 or HSV-2 challenge as compared to the mock vaccinated challenge group. T-cell memory immune responses to specific glycoprotein B (gB) and glycoprotein D (gD) peptide epitopes were detectable at 7 months post vaccination.ConclusionsThese results suggest that the highly attenuated, non-neurotropic gK-null virus may be used as an effective vaccine to protect against both virulent HSV-1 and HSV-2 genital infections and induce lasting immune responses.

Application of shRNA-containing herpes simplex virus type 1 (HSV-1)-based gene therapy for HSV-2-induced genital herpes

HSV-1-based vectors have been widely used to achieve targeted delivery of genes into the nervous system. In the current study, we aim to use shRNA-containing HSV-1-based gene delivery system for the therapy of HSV-2 infection. Guinea pigs were infected intravaginally with HSV-2 and scored daily for 100 days for the severity of vaginal disease. HSV-2 shRNA-containing HSV-1 was applied intravaginally daily between 8 and 14 days after HSV-2 challenge. Delivery of HSV-2 shRNA-containing HSV-1 had no effect on the onset of disease and acute virus shedding in animals, but resulted in a significant reduction in both the cumulative recurrent lesion days and the number of days with recurrent disease. Around half of the animals in the HSV-2 shRNA group did not develop recurrent disease 100 days post HSV-2 infection. In conclusion, HSV-2 shRNA-containing HSV-1 particles are effective in reducing the recurrence of genital herpes caused by HSV-2.

The efficacy of HSV-2 vaccines based on gD and gB is enhanced by the addition of ICP27

DNA vaccines expressing HSV-2 gD, gB, ICP27, VP22 and VP13/14 were shown to be immunogenic in mice; gD and gB elicited neutralising antibody, and all five antigens induced T cell responses measured by IFNγ ELISPOT. In murine HSV-2 challenge studies, gD and gB provided moderate to high levels of protection while ICP27 provided a lower level of protection depending on the model (intravaginal or intranasal) and the challenge dose. Combining vaccines expressing gB or gD with vaccines expressing ICP27 provided greater protection than any antigen alone. We conclude that the addition of ICP27 to enhance the anti-viral T cell response can improve the efficacy of gD- and gB-based vaccines.

A Vaxfectin®-adjuvanted HSV-2 plasmid DNA vaccine is effective for prophylactic and therapeutic use in the guinea pig model of genital herpes

Here we describe studies in the guinea pig model of genital herpes to evaluate a novel plasmid DNA (pDNA) vaccine encoding the HSV-2 glycoprotein D and UL46 and UL47 genes encoding tegument proteins VP11/12 and VP 13/14 (gD2/UL46/UL47), formulated with a cationic lipid-based adjuvant Vaxfectin®. Prophylactic immunization with Vaxfectin®-gD2/UL46/UL47 significantly reduced viral replication in the genital tract, provided complete protection against both primary and recurrent genital skin disease following intravaginal HSV-2 challenge, and significantly reduced latent HSV-2 DNA in the dorsal root ganglia compared to controls. We also examined the impact of therapeutic immunization of HSV-2 infected animals. Here, Vaxfectin®-gD2/UL46/UL47 immunization significantly reduced both the frequency of recurrent disease and viral shedding into the genital tract compared to controls. This novel adjuvanted pDNA vaccine has demonstrated both prophylactic and therapeutic efficacy in the guinea pig model of genital herpes and warrants further development.