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Medicine » Infectious Diseases » "Herpesviridae", book edited by Jozsef Ongradi, ISBN 978-953-51-2611-9, Print ISBN 978-953-51-2610-2, Published: September 7, 2016 under CC BY 3.0 license. © The Author(s).

Chapter 7

Herpes Simplex Virus 1 and 2 Vaccine Design: What can we Learn from the Past?

By Vladimíra Ďurmanová, Marian Adamkov and Július Rajčáni
DOI: 10.5772/64447

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Herpes Simplex Virus 1 and 2 Vaccine Design: What can we Learn from the Past?

Vladimíra Ďurmanová1, Marian Adamkov2 and Július Rajčáni3
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This chapter is devoted to the topics of not yet marketed HSV vaccine, which is still in the focus of interest, especially from the point of immunotherapeutic use. To understand the principles of vaccination strategies (prophylactic and/or immunotherapeutic), the pathogenesis of herpes simplex virus 1 (HSV-1) and/or HSV-2 infections in animal models is briefly outlined. Even when both herpesviruses may spread via bloodstream, which is especially true in the immunocompromised host, the main route of their transmission is along peripheral nerves. Both viruses establish latency in ganglion cells, and after reactivation, they spread along axons back to the site of primary infection. Since neither the establishment of latency nor its reactivation can be fully controlled by virus-neutralizing antibodies, the outcome of immune response greatly depends on the activity of cytotoxic CD8+ T lymphocytes. The majority of important antigenic epitopes is located in envelope glycoproteins (such as gB, gD, gE, gC and gG) that are related to virus adsorption and penetration into susceptible cells. The HSV-1 and/or HSV-2 experimental vaccines designed so far were either purified virion products derived from infected cells (subunit vaccines), purified recombinant immunogenic herpes simplex virus HSV-coded proteins (especially gD), and/or attenuated live viruses lacking some of virulence tools (such as gH and/or gE). We bring a comprehensive overview of the efficacy of experimental HSV-1/HSV-2 vaccines and discuss our own data. In conclusion, we believe in the continued demand of HSV-1 and HSV-2 vaccines, at least for their immunotherapeutic use, suggesting unified evaluation criteria for clinical trials to reach consent at their interpretation.

Keywords: herpes simplex virus 1 and herpes simplex virus 2, candidate vaccine, clinical trials, efficacy in animals, humoral and cell-mediated immune response, antigenic epitopes

1. Introduction

This review will be devoted to the topics of not yet marketed HSV vaccine(s), which design is still in the focus of interest, especially from the point of view of immunotherapeutic use. The human herpesvirus 1 (common name herpes simplex virus 1, HSV-1) is one of the first human viruses discovered [1] and belongs among the most intensively investigated viruses. In the span of the last 45 years, the PubMed portal has registered nearly 11,000 papers devoted to various aspects of HSV-1/HSV-2, starting from virus structure, continued by the molecular mechanisms of lytic replication versus latency maintenance, through the virus spread in the body and the eliciting of different forms of immune response, not omitting many clinical and epidemiological studies. In the second half of the last century, two subtypes of herpes simplex virus (HSV) have been described [2] and were designated HSV type 1 and HSV type 2 [3]. While the former (HSV type 1) has been predominantly isolated from the orofacial area and upper respiratory airways, the latter was believed to infect the urogenital tract and occasionally the newborn. According to recent classification, the HSV-2 represents a distinct species of the Simplexvirus genus, which along with the Varicellovirus genus belongs to subfamily Herpesvirinae of the Herpesviridae family. Nevertheless, both species are closely related, since they differ only in a few antigenic domains (and/or epitopes) located in the envelope glycoproteins, namely, in an entirely distinct gG and/or in the partially unrelated gC [4].

2. The molecular biology of HSV: virion structure, lytic replication and latency

The HSV virion consists of four elements: (a) a core containing the viral DNA (vDNA), (b) an icosahedral capsid surrounding the core, (c) an unstructured proteinaceous layer called the tegument that surrounds the capsid, and (d) an outer lipid bilayer envelope exhibiting spikes on its surface. The core contains the double-stranded (ds) DNA (dsDNA) genome wrapped as a toroid or spool in a liquid crystalline state. A small fraction of the virion DNA may be circular, but the bulk of packaged HSV DNA is linear and double-stranded. The enveloped virion is a spherical particle with an average diameter of 186 nm, which might extend to 225 nm with the spikes included, while the internal capsid has a constant diameter of 110 nm (reviewed at [5]). The HSV virion contains more than 30 proteins that were designated as virion polypeptides (VPs). Out of these approximately known and additional 10 suspected virion proteins (VPs), at least 11 are located within the envelope at the surface of virion (predominantly accessible to antibody), from which at least 10 are glycosylated (reviewed at [6]).

The HSV genes were classified into at least three general kinetic classes: alpha or immediate-early (IE), beta or early (E), and gamma or late (L) genes. The IE mRNAs are transcribed by the help of a transcription-inducing cofactor present in the virion tegument, also called alpha-TIF/VP16. In brief, the tegument protein VP16 is becoming a part of a tripartite complex comprised of octamer-binding protein 1 (Oct-1) and host cell factor 1 (HCF-1). At least two of the IE proteins (ICP0 and ICP4; Figure 1) are transactivators needed for initiation of transcription of the E/beta mRNAs (the first checkpoint) and later also for transcription of gamma/L mRNAs. The latter are mainly structural virion proteins in contrast to E transcripts specifying nonstructural polypeptides. The structural proteins are transcribed in two waves, called gamma1/L and gamma2/L. The former occurs more closely related with the E transcription phase (second checkpoint) and requires predominantly ICP4. The expression of beta polypeptides as well as that of group gamma1/L genes requires ICP4 and at low multiplicities of infection also ICP0. The expression of gamma2/L genes, controlled at the third checkpoint, requires ongoing DNA synthesis as well as the presence of ICP4 and ICP0 [7].


Figure 1.

The IE genes and positions of their ORFs in the HSV genome.

The HSV genome consists of two covalently linked sequence components, designated as L (long) and S (short). Each component consists of unique (U) sequences bracketed by inverted repeats (IR) [6].

The entry of HSV into cells involves the interaction of at least five virion surface glycoproteins (gB, gC, gD, gH and gL) with several receptors on the cell surface following fusion of the envelope with a cellular membrane. The HSV glycoproteins along with other proteins important for pathogenicity are listed in Table 1. The first step in the process of entry is the binding of virions to glycosaminoglycans (GAGs) on the cell surface. The interaction of virions with GAGs is mediated by two glycoproteins: gC and gB. The next step in virus entry consists of the interaction of gD with its receptors and execution of fusion between the envelope and the cellular membrane by the heterodimer gH/gL and also by gB. The interaction of gD with its receptors has been extensively studied in several laboratories. The consensus is that, for entry, gD interacts with one of three natural receptors (reviewed at [8]). The receptors are nectins, a protein-designated herpesvirus entry mediator (HVEM) and a selected form of 3-O-sulfated heparan sulfate (3-OS HS). Nectins are intracellular adhesion molecules expressed on epithelial and neural cells and are members of the extended immunoglobulin (Ig) family. Earlier studies showed that gD of HSV-1 interacts with the amino-terminal V1 domain of nectin 1. The wild-type HSV-2 can enter susceptible cells via an alternative receptor, namely, nectin 2. HVEM is a member of the extended tumor necrosis factor receptor (TNFR) family, expressed mainly on T lymphocytes but occasionally also on natural killer (NK) cells. Its natural ligand is LIGHT (homologous to lymphotoxins, exhibits inducible expression), which is constitutively expressed on T and natural killer (NK) cells and appears to be a regulator of mucosal immune system. The LIGHT receptor competes with HVEM for the gD (reviewed at [9]). The role of gD with respect to gB and gH/gL is to recruit and position them properly to enable interaction with GAGs and with the lipid bilayers of the cellular membranes [10].

GeneProtein Function
RL1γ34.5L protein, cofactor for ribosomal translation
RL2ICP0IE protein, spliced, three exons, a RING finger motif, acts as a nonspecific transactivator of any beta and gamma gene
UL1gLLate glycoprotein, forms the gH/gL complex, syn mutation
UL5E class protein, part of the helicase/primase complex
UL9E/L class, ori-binding protein, cooperates with ICP8 at initiation of vDNA replication
UL10gML class, glycosylated polypeptide, interacts with gN, participates in exocytosis and in cell-to-cell spread
UL18VP24L class, capsid component, participates in triplex formation
UL19VP5L class protein, the major capsid component (149/150K)
UL21L class tegument protein weakly capsid-associated, binds to microtubules at axonal transport
UL22gHL class, glycosylated protein, essential for penetration to cells, needs complexing with gL
UL23TKE class, tegument protein, thymidine kinase, virulence factor
UL26L class, “scaffolding” capsid protein, the N-terminus required for virion assembly
UL27gBL class glycoprotein, essential for adsorption and membrane fusion, syn3 locus
UL29ICP8E class, ssDNA binding, required for vDNA synthesis, keeping the DNA fork apart
E/L class protein, DNA polymerase (elongation enzyme), virulence factor
UL35VP26L class virion protein, hexon component, also termed NC7
L class, large tegument protein, important for egress through cytoplasm and re-envelopment
UL38VP19CL class capsid protein, triplet component with VP23 (1:2), connects the hexons and pentons
UL39RRE class protein, large subunit of RR, membrane-anchored protein kinase
UL40RRE class protein, the small subunit of RR
UL41 vhs L class, tegument protein, interferes with the host cell proteosynthesis, virulence factor
UL44gCL class, glycoprotein, reacts with GAG on cell surface, binds C3 and/or C5, virulence factor
L class tegument protein, alpha-transinducing factor
UL49.5 gNL class membrane-associated small glycoprotein, complexes with gM
UL53gKL class glycoprotein (40K), localizes to Golgi and ER, involved in egress, syn1 locus
IE class protein, blocks cellular mRNA transport to cytoplasm
RS1ICP4IE class protein, transactivator for E and L promoters
US1ICP22IE class regulatory protein, essential in animal experiments, cellular cyclin A and B degradation
US4gGL class, glycosylated protein and envelope component, HSV-2 specific
US6gDE/L class, glycosylated protein, 56K, adsorption to nectin 1 and HVEM protein receptors, essential for entry to cells, neural uptake
US7gIL class glycoprotein, forms heterodimer with gE, the complex facilitates cell-to-cell spread, neural uptake
US8gEL class glycoprotein, complexes with gI, essential for neural uptake
US12ICP47IE class protein, blocks the transport of viral antigenic epitopes
LATThree categories of RNAs (8.5 kb, 2.0, and 1.5 kb), the small LATs terminate antisense to ICP0 ORF. Latency regulator

Table 1.

HSV-1 genes and corresponding proteins regarded for importance in virulencea.

[i] - Abbreviations: vDNA = viral DNA; ICP = infected cell protein; VP = virion protein (structural); IE = immediate-early kinetics (class); E = early kinetics (class); L = late kinetics (class); K = 1,000 kDa (Mr); RR = ribonucleotide reductase; vhs = virion host shutoff; LAT = latency-associated transcript(s); RNA molecules without capping, do not interact with ribosomes; UL = unique long (DNA segment); US = unique short; RL = repeat long; RS = repeat short; HVEM = herpes virus entry mediator)

[ii] - aModified according to Roizman et al. [6].

The gB belongs to class III fusion protein that after GAG interaction facilitates fusion of the virion envelope bilayer with the cellular membrane. A recent study showed that gB can also bind to the paired immunoglobulin-like type 2 receptor-α (PILRα) to trigger viral fusion in certain cell types; however, the precise role of this interaction in viral entry remains to be determined [11]. The gH and gL appear to form a tight complex, since neither protein is stable without the other. In contrast to gB, the structural studies of gH/gL showed no homology with fusion domains of other viral glycoproteins. The exact role of gH/gL has not been determined; however, one hypothesis is that the interaction of gD with its receptors changes the conformation of gH/gL, which in turn induces gB to adopt its fusion conformation [12]. Even that gE had been regarded for not essential in cell culture, it has been shown very important at entering nerve endings (neural uptake) and subsequent neural spread [13].

The latency of HSV can be characterized by circularized state of the HSV genome in the absence of IE and/or E transcription, especially lacking the expression of the two most important beta/gamma transcription activators, namely, ICP0 and ICP4 (reviewed at [14]). We pointed at the possibility that the IE block in question might be “leaky” since small amounts of ICP4 mRNA could be found in the non-cultured ganglion explants which would later on yield infectious virus [15]. Overwhelming literature deals with the hypotheses that the small latency-associated transcripts RNA (LATs), RNA molecules expressed during latency contribute to the silencing of ICP0 mRNA transcription as well as to the maintenance of latency [16]. It has been shown that LAT can reduce the expression of viral genes and suppress HSV replication in cultured cells. In addition, LAT probably protects the HSV DNA carrying neuron from apoptosis. The anti-apoptosis activity of LAT has been independently confirmed in tissue culture and in the mouse ocular model [17]. Further data suggest the participation of LAT in reactivation of the latent genome [18, 19].

3. Pathogenesis of HSV-1 and/or HSV-2 infections

The crucial event in HSV pathogenesis is, when the virus, which has reached the pseudounipolar neuron of the regional sensory ganglion via the quick axonal transmission, may but need not undergo productive replication. Thus, latency can be established from the very beginning, that is, in the absence of any transcription provided by the virus-coded alpha-transinducing factor (alpha-TIF). As described by Efstathiou et al. [20], the latent genome resides in the nuclei of neurons in the form of a circularized nonintegrated plasmid-like structure. This structure survives within the carrier neuron. the carrier neuron, so that neural cells harbor the viral DNA for their lifetime. The tegument-bound alpha-transinducing factor (alpha-TIF) might be “lost” during intra-axonal transport [21]. Some axons might be as long as 500 mm (as a rule about 100 mm), while the minimum speed for HSV capsid transmission is about 1–2 mm/h. The quick axonal transport along neurotubules is provided by the dynein molecule, which binds the capsid component UL21. The environment within sensory neurons favors the establishment of latency rather than productive virus replication also because the neuronal transcription cofactors such as Oct-2 and/or Brn-3 repress the IE promoters [22, 23].

During reactivation, the blocked IE checkpoint of virus replication must be overcome especially by overwhelming expression of ICP0 protein in order to achieve productive virus replication and infectious HSV production [24]. The reactivation process can be hampered by two different mechanisms. One depends on viral genome products interacting with cellular cofactors of transcription within neurons. It may not be excluded that due to the existence of at least two activation checkpoints, controlling the expression gamma1 or E/L proteins (glycoproteins) and the gamma2/L proteins (glycoproteins), also incomplete virus particles may be formed. The E and/or E/L proteins may be synthesized in the absence of complete virion formation, or assembly of virions with defective vDNA occurs. However, the presence of virus-coded antigens (even in the absence of infectious virus particles) could induce a potent immune response (including the accumulation of cytotoxic T cells in the vicinity of HSV carrier neurons). It should be also mentioned that the IE protein ICP47 acts by immune evasion. Alternative effects inhibiting the antiviral defense can be exerted by gC [25], which binds complement, and by the gE molecule, which binds the Fc fragment of immunoglobulins.

If reactivated virus reaches the peripheral tissues by retrograde axonal transport (skin squamous epithelium, non-cornified squamous epithelium of mouth mucous membrane, corneal epithelium), then it starts to replicate and causes inapparent virus shedding or blister formation. A nice example for creating favorable conditions of virus replication at peripheral skin was described in the ear model [26, 27]. These investigators found that prostaglandins produced after skin trauma or UV light irradiation would enhance the replication of recurrent HSV-1. As shown by Walz et al. [28], neurectomy of the trigeminal nerve root reactivated the latent HSV harbored within ganglion cells. The round trip of reactivated virus usually ends with reinfection of additional neurons within the regional sensory ganglion. Less frequently, the retrograde axonal transport may continue in centripetal direction, that is, to brain stem. Furthermore, the central nervous system (CNS) may become infected via bloodstream (in newborn) or along the olfactory route (reviewed at [29]).

The different manifestations of clinical disease reflect the above-described mechanisms of HSV spread in the body. Essentially, HSV-1 can induce the acute primoinfection (gingivostomatitis) and recurrent disease (classical labial herpes). Alternatively, the ocular herpes, often manifested as herpes keratitis, occurs rather as recurrent disease than primoinfection. In newborn as well as in the immunocompromised host and/or in the case of local inflammation in the skin (e.g., due to allergic manifestations such as atopic eczema), the HSV might cause generalized skin disease or even viral sepsis (i.e., the hepatoadrenal necrosis in newborn or severe meningoencephalitis). In newborn, perinatal infection causes an acute dissemination of HSV-2 via bloodstream. Meningitis and meningoencephalitis of neonates, similarly as various forms of genital infection of man and women, are mainly HSV-2-related. In women, the outer but also the inner genital tract (vagina or cervical mucosa covered with non-cornified squamous epithelium) might become infected. In the skin as well as at the mucous membranes covered by squamous epithelium, both HSV-1 and HSV-2 species replicate within the lower and/or medium squamous cell layers causing blister formation [30].

4. Mechanisms of the immune response to HSV-1 or HSV-2

The immune response to HSV-1 and/or HSV-2 is induced in an early and a late phase. During early phase, the nonspecific antiviral mechanisms are activated, while in the late phase, the HSV-specific reaction is mobilized. The specific immune response culminates by recruiting the CD8+ cytotoxic T lymphocytes important for extracellular latency control, in contrast to the intracellular latency control which has been discussed above. The specific cell-mediated immune response may begin in the regional lymph nodes draining the virus inoculation site (both T-lymphocyte lines are engaged) as well as in the trigeminal ganglia which harbor the latent virus (mainly cytotoxic CD8+ T cells take part).

4.1. The innate immune response

The innate antiviral immunity to HSV-1 and/or HSV-2 is provided by alpha/beta type I interferons (IFNs) as well as by activation of natural killer (NK) cells. The role of type I IFNs in anti-HSV defense was demonstrated in knockout mice deficient in type I IFN production [31, 32]. The type I IFN release is induced by virus constituents called pathogen-associated molecular patterns (PAMPs) interacting with the corresponding pattern recognition receptors (PRRs) at the surface of responding cells. The best known examples are the Toll-like receptors (TLR). For example, the envelope glycoprotein D molecule of the HSV-1 interacts with the TLR-2 receptor on the target cell membrane [33], while the TLR-9 receptor binds to non-methylated CpG motifs of the vDNA present in the nuclei of infected cells [34]. It has been also shown that children with TLR-3 defects are more susceptible to herpes encephalitis [35, 36]. Both types I of IFNs act on the receptors in neighboring cells eliciting the synthesis of antiviral substances such as the 2′,5′-oligoadenylate synthase, RNase L, and/or RNA-dependent protein kinase. The type I IFN also inhibits cell proliferation and enhances the activation of NK cells as well as the expression of MHC class I molecules, but downregulates MHC class II expression. The type III IFNs, also designated as IFN-λ1, IFN-λ2 and IFN-λ3 (IL-29, IL-28A, and IL-28B), have been recently characterized; alike to classical type I IFNs, also IFN-λ induces production of antiviral substances in the infected cells [37]. The participation of type III IFNs in inhibition of HSV replication has been confirmed in vitro [38, 39].

In addition to type I IFN formation, the activation of NK cells represents another important arm of nonspecific immune response to HSV-1 and HSV-2 infection [40]. The nonspecific killers belong to the group of class I innate lymphoid cells. Morphologically, they belong to large granular lymphocytes (LGLs), which cytoplasm contains azurophilic granules with perforins and granzymes A and B. They exert cytolytic activities helpful at the elimination of infected cells. The perforins create a transmembrane channel piercing the cell membrane, while granzymes activate the apoptosis of target cells. The NK cells also produce IFN-gamma, which promotes their activation along with the activation of macrophages. In association with this, enhanced death rates were noted in NK-deficient mice infected with HSV [41].

The elimination of HSV-1 and/or HSV-2 infected cells at the portal of entry is mediated by additional immune cells such as neutrophils, dendritic cells (DCs), and macrophages. The neutrophils secrete tumor necrosis factor (TNF) which induces apoptosis of infected cells via caspase-8 activation [42]. Blood monocytes when entering the connective and/or other tissues are altered into macrophages able to engulf the extracellular virus particles but also residues of apoptotic cells. The active macrophages belong to the group of antigen-presenting cells (APCs) that present the external immunogenic peptides to T lymphocytes. This process called antigen presentation launches the specific immune response. Activated macrophages also release several pro-inflammatory cytokines like TNF and IL-6 (participates in B-cell activation), type I IFNs, and several chemokines such as RANTES regulated upon activation normal T-cell expressed and secreted. Finally, nitric oxide (NO) and other substances are produced by macrophages. It has been demonstrated in experiments in vitro that the replication of HSV-1 in infected cells may be considerably inhibited in the presence of NO released from macrophages [43]. It should be mentioned here that IFN-gamma (belongs to type II IFNs) is in fact a cytokine released from activated NK cells but also produced by the helper CD4+ T lymphocytes as well as by cytotoxic CD8+ T lymphocytes [44]. The IFN-gamma considerably promotes function of activating macrophages and NK cells, in which it induces a production of cytolytic substances. Taken together, both NK cells and macrophages represent an important first-line defense at the early stage of HSV infection, that is, before the onset of specific immune response.

As already mentioned above, important immune cells participating in the initiation of antigen processing are the dendritic cells (DCs). They represent a relatively heterogenous cell population, from which the most active in antiviral defense are the myeloid (conventional) and the plasmacytoid DCs (pDCs), both derived from the myeloid progenitor cells. The myeloid DCs act as antigen-presenting cells (APCs), which carry the processed antigenic peptides into lymph nodes, where the peptides in question are presented to the T lymphocytes. Based on this presentation event, the specific immune response is induced (activation of T and B cells). In DC-depleted mice infected with HSV-1, encephalitis developed with a significantly higher frequency as compared to conventional mice [45]. The plasmacytoid DCs under physiological conditions participate in the development of peripheral immune tolerance. Upon HSV infection, plasmacytoid DCs start to produce high amounts of IFN-alpha, which not only inhibits the virus replication in the otherwise surrounding susceptible cells but also activates lymphocytes and additional DCs. The HSV dsDNA binds to TLR-9 of the plasmacytoid DC which initiates IFN-alpha production [46].

4.2. The acquired immune response

The acquired immune response encounters the activation of both T-lymphocyte lines, namely, those differentiated either in direction of CD4+ or CD8+ T cells. The acquired immune response then is triggered by the activation of T-cell receptors (TCRs), which recognize the HLA I/CD8 and/or the HLA II/CD4 bound antigenic peptides. While the former T cells are involved in the destruction and elimination of HSV-infected cells, the latter acts as helper T cells at inducing the specific antibody production by B cells. At primary infection, the HSV-1 or HSV-2 particles are engulfed by DCs, which move the viral peptides to the regional lymph nodes, where the naïve (unprimed) lymphocytes get first stimulated. The presentation of exogenous viral peptides is achieved by means of the HLA class II molecules, which are recognized by the TCR of the CD4+ T lymphocytes. In contrast, the immunogenic peptides of the newly (de novo) synthesized HSV proteins are presented by HLA class I molecules at the surface of infected cells where they interact with the TCR of cytotoxic CD8+ T lymphocytes [47]. In certain extent, the so-called cross presentation occurs, at which the exogenous viral antigens are binding into the HLA class I molecules that are recognized by the CD8+ T lymphocytes [48]. The activated helper CD4+ T cells release cytokines such as IL-2 (inducing T-lymphocyte proliferation) and IL-4, IL-5 and IL-6 (promoting the differentiation of B cells into plasma cells).

The plasma cells synthesize the specific virus-neutralizing antibodies reacting mainly with the envelope glycoproteins such as gB and gD. The antibodies may also activate complement; on the other hand, the gC molecule of the HSV-1 envelope binds C3b and in less extent also C5b reducing the availability of complement components for virus neutralization. The low-pathogenic HSV-1 gC minus strains may become pathogenic as was confirmed in a C3 knockout murine model [49]. The HSV-1 antibodies belong to IgM as well as IgG class; the HSV-2 antibodies may be also of the secretory IgA class; the latter participates in virus clearance at the genital tract mucosa [50]. Even though the serum antibodies are of importance at the acute phase of infection, they may not fully eliminate the HSV since it invades the nerve endings and spreads to regional sensory ganglia before the development of antibody response. Some studies have shown that passive immunization with immune serum did not prevent latency after genital infection with HSV-2 [51, 52, 53]. Since the latent HSV (episomal vDNA) is frequently harbored in neural tissue for lifelong, the presence of HSV antibodies in the serum of healthy subjects may be interpreted as infectious immunity.

The cytotoxic CD8+ T lymphocytes represent the mainstream of the specific immune response providing clearance of HSV-1- or HSV-2-infected cells from the body. The activation of cytotoxic CD8+ T cells takes place in the regional lymph nodes by means of viral antigenic peptides presented by HLA class I molecules expressed at the surface of DCs. The IL-2 released from helper T cells acts on the precursor cytotoxic T lymphocytes which differentiate into mature cytotoxic CD8+ T cells. These mature CD8+ T cells accumulate at the peripheral virus inoculation site, where they eliminate the HSV-infected cells. The activated cytotoxic T lymphocytes release substances such as perforins, granzyme and granulysin, which destroy the infected target cell and/or induce apoptosis acting on their FAS receptors. They also release cytokines such as IFN-gamma and the tumor necrosis factor (TNF). IFN-gamma has a multiple effect, since it enhances the expression of HLA class I as well as HLA class II molecules [54] and also induces the expression of antiviral substances such as protein kinase R (PKR). This substance causes inhibition of translation of many viral but also cellular proteins. The increased TNF production also enhances the number and activity of HLA class I molecules.

The clearance of HSV-2 from peripheral tissues such as genital mucosa may be also provided by CD4+ T cells. Transgenic mice, which revealed defects in their CD8+ T-lymphocyte activity, have been still well protected against a lethal dose of HSV-2. In contrast, mice with depleted CD4+ T lymphocytes showed slower clearance and less protection against HSV-2 [5557]. It can be concluded that the clearance of HSV-2 from genital mucosa requires cooperation of both the helper and cytotoxic T lymphocytes. Both T-cell populations can be found in the skin, in the genital and oral mucosa, in the ocular tissue, and also in the trigeminal ganglia surrounding the infected neurons [5860].

After healing of the acute phase of HSV infection, about 0.1–1% of memory T cells, which may remain within the circulation, are still able to recognize the HSV antigenic epitopes. These epitopes being recognized by CD4+ and/or CD8+ lymphocytes were not all exactly mapped yet, but until now, 22 of them were defined. The HSV-specific epitopes can be found mainly among VPs (structural envelope glycoproteins, tegument proteins and capsid polypeptides), but are also present on the ICPs, that is, on nonstructural enzymes and regulatory proteins. The CD4+ T lymphocytes generated against the HSV-2-specified structural proteins recognize the epitopes of the RR1/UL39 polypeptide, the epitopes of UL46 tegument protein, and several ones on glycoproteins gD and gB [61].

The CD8+ T lymphocytes, surrounding the neurons of the regional sensory ganglia of mice, rabbits, and humans in which viral latency had been established [6264], are believed to control reactivation [65]. It was demonstrated, for example, that the ICP4/145K IE protein can be digested by granzyme B, which would prevent virus reactivation [66]. The accumulated CD8+ T cells produce IFN-gamma as well, inhibiting the expression of ICP0/110K IE protein [67].

The most intriguing still unexplained question is why do the CD8+ T lymphocytes accumulated in the neighborhood of latent HSV-harboring neurons, since the expression of IE and E polypeptides is widely hampered in them. Interestingly enough, the great majority of CD8+ T lymphocytes in the trigeminal ganglion of mice in which latency had been established possess a TCR which reacts with gB epitope at aa 498–505, while a smaller proportion of them may be stimulated with RR1 epitope aa 822–829. The reactivity of the other T cells was not identified [68, 69]. In human trigeminal ganglia, both the CD4+ and CD8+ T cells were described that react with many antigenic epitopes present in the IE polypeptides ICP0 and ICP4, in the E polypeptides such as thymidine kinase (TK) and RRl, as well as with antigenic domains from structural proteins such as the tegument proteins VP11/12 and VP13/14 and envelope glycoproteins gB, gK, and gL, even though an exact identification of all antigenic epitopes in the HSV proteins was not done yet [70, 71].

The phenotype of CD8+ T lymphocytes which can accumulate within the ganglia was not characterized. The permanently present cytotoxic T lymphocytes might belong to the population of so-called tissue-resident memory T lymphocytes (TRM) or to the population referred as T-effector memory cells (TEM cells). Additional markers such as CD44, CD69, CD62L, and CCR7 were described at their surface, but their full definition is still the matter of investigation [72].

5. Survey of experimental HSV-1 and HSV-2 vaccines and their design

Though the first vaccines against HSV-1 and/or HSV-2 were prepared in the sixties of last century [73], no accepted and fully efficient HSV vaccine is available till now. The main problem is the incomplete understanding of the role of cytotoxic CD8+ T cells in the maintenance and/or of elimination neurons carrying the silenced HSV DNA. Another serious problem is the production of immune evasion polypeptides (reviewed at [74]). The optimal HSV vaccine should overcome the ICP47-mediated interference with TAP, which transports the viral peptides inside ER to bind with HLA molecules. Finally, the last but not negligible problem is that at least some of over 20 immunogenic epitopes, which are distributed within about 80 HSV proteins (both structural and nonstructural), should participate in the vaccine but in an optimal composition. An effective HSV vaccine might differ depending of its purpose, which could be either preventive or immunotherapeutic. An ideal vaccine should not allow the acute disease to develop, and it should be able to prevent the establishment of latency. These goals, especially the latter, are difficult to achieve. The minimum effect provided by an immunotherapeutic vaccine would be to minimize the extent of recurrent lesion originating from the reactivation of the latent viral genome. As described above, the cytotoxic T lymphocytes accumulating in vicinity of HSV DNA carrier cells in the regional sensory ganglion seem of great importance when keeping a state of equilibrium probably inhibiting the transcription of IE genes by an unknown mechanism. One could estimate that in the case of the inefficiency and/or complete failure of the function of cytotoxic T-cell virus, reactivation and subsequent retrograde axonal transport of virions occur. In this section, we describe the various HSV vaccines which have been used under experimental conditions, while the next paragraph will describe their efficacy in human volunteers.

5.1. Inactivated virion and subunit vaccines

The inactivated HSV-1 and/or HSV-2 vaccines consist of infected cell extracts, which have been introduced in the 1980s of last century, partially purified and inactivated either by UV light, by thermal treatment, or with formalin [7577]. The efficacy of these first vaccines was tested mainly in mice, but some of them were used also in human trials (see Section 6). Especially in animal models, a good protective effect against acute virus challenge has been found not to prevent the establishment of latency. However, when testing the effect of a virion-free HSV vaccine prepared by extraction of infected LEP cells with Nonidet P-40 [78] in the rabbit corneal model, Rajčáni et al. [79] concluded that even though vaccination prior to infection did not fully prevent latency, it considerably reduced the number of regional sensory (trigeminal) ganglion cells (neurons), which became the HSV genome carriers. Recently, a formalin-inactivated HSV-2 (FI-HSV2) vaccine mixed with Al(OH)3 adjuvant was tested in combination with monophosphoryl lipid A (MPL) [80]. The latter authors showed that the vaccine protected mice against local vaginal challenge with HSV-2 as well as reduced the extent of latency established in the sensory ganglia of lumbosacral nerve roots.

The subunit vaccines were mainly glycoprotein mixes purified on lectins (e.g., lentil lectin) showing high affinity to the HSV virion envelope antigens. The dominant protective antigens among the 11 HSV glycoproteins are gB and gD, which possess important immunogenic epitopes. The abovementioned glycoproteins elicit virus-neutralizing antibodies, antibodies participating in the antibody-dependent cellular cytotoxic (ADCC) response, and they also activate the T lymphocytes. Several reports described the efficacy of the subunit vaccines of various purity based on either HSV-1 and/or HSV-2 envelope glycoproteins [8184]. For example, the subunit HSV-1 vaccine (strain HSZP Immuno) had been prepared from chick embryo cells infected with the low-virulent HSZP strain [85]. Tested in cooperation with the Research and Development Department of the former Immuno AG Company in Vienna, the infected cell extract in question was purified on lentil lectin to obtain a glycoprotein mix containing at least four envelope glycoproteins (gB, gC, gD and gG). This subunit vaccine was immunogenic and protective in mice as well as rabbits and showed at least partial cross protection in the HSV-2-challenged guinea pigs infected by the vaginal route.

5.2. Recombinant HSV-1 and HSV-2 vaccines

The next step of HSV vaccine development proceeded from purified (nearly cell DNA-free) subunit vaccines to those composed of recombinant HSV-1/HSV-2 polypeptides [8690]. Great majority of the recombinant vaccines contained the gB and/or gD polypeptides, and their efficacy was done in mice. The first purified glycoprotein vaccines (gB1 and gD1) were prepared on immunoaffinity columns (with specific-bound MoAbs); such vaccines were found to protect mice against intracerebral challenge with HSV-1 [9193]. Manservigi et al. [94] expressed the gB ectodomain in mammalian cells. After immunization with the purified gB fragment, the mice developed virus-neutralizing antibodies not only against HSV-1 but also cross-reacting antibodies to HSV-2. In addition, the authors also showed that the animals were resistant to the challenge with a lethal HSV dose. Later on, the recombinant proteins were prepared by transfection of plasmids (carrying the gD and/or gB ORFs) into competent Escherichia coli cells, which expressed the corresponding not glycosylated polypeptides [90, 95100]. Recombinant glycoproteins (including gC) were also prepared in insect and/or mammalian cells, in which the recombinant products could be glycosylated [101].

Immunization with the recombinant gD1 or gD2 alone, or in combination with the gB protected mice against infectious virus challenge given by intraperitoneal or subcutaneous routes and in the case of gD1 vaccination cross protection to HSV-2 was observed as well. Immunization with gD alone was more effective than that with gB alone [87], while immunity provided by gC alone was negligible [95]. In contrast, very high effect was achieved following immunization with the mix of gB1, gD1, and gE1 glycoproteins in combination with the Al(OH)3 adjuvant including the cross protection of guinea pigs challenged with HSV-2 at genital route [99]. A trivalent vaccine containing gD2, gC2 and gE2 prepared in the baculovirus expression system was more immunogenic for mice than the subunit vaccine containing gC2 and gD2 [102]. Furthermore, Awasthi et al. [103] immunized the guinea pigs with recombinant proteins gC2 and gD2 prepared in a baculovirus system (which activated antibody production), with the adenovirus vector carrying the VP5 capsid protein and with the tegument proteins VP13 and VP14 (which activated the T-cell-mediated response). However, no significant difference was detected, which confirmed that the glycoprotein mix gD2/gB2 was sufficient enough.

The alum adjuvant in combination with the recombinant gD polypeptide activates the Th2-type humoral response and therefore may be less suitable for immunotherapeutic use, for which the Th1-type response is essential. To stimulate the Th1 response predominantly, cytokines as adjuvants were tested, such as IL-2 with good results after corneal challenge with the virulent HSV-1 CHR3 strain [104]. The so-called immune-stimulating lipid complexes (ISCOM) combined with HSV-2 immunogenic polypeptides induced both Th1- and Th2-type responses when showing a highly neutralizing antibody response along with the production of IL-2, IFN-gamma, and TNF [105]. Similar strong potentiating effect was observed at immunization with gD2 polypeptide adjuvanted with modified lipid A (AS04) [98]. Furthermore, when using the gD2 fusion polypeptide along with MF59 (squalene) as adjuvant, the local mucosa immune response could neither be stimulated properly, nor the latency reactivation prevented [106]. There is worth mentioning that the so-called autoimmune/inflammatory syndrome induced by adjuvants (ASIA) is an inflammatory syndrome associated with certain adjuvants (such as MF59/squalene); the latter potentiates pathological autoimmune reactions, which may be evident in the case of prophylactic mass immunization campaigns [107].

In contrast to T-lymphocyte stimulation, the local IgA secretion can be enhanced by mistletoe lectins such as ML-1, ML-2, and/or ML-3, the application of which in combination with the gD2 polypeptide elicited a good secretory IgA response at intranasal challenge in mice [108]. The authors also used non-ionized liposome like particles carrying the gB ectodomain and additional polylysine-rich polypeptides. Intranasal application of the product in question provided protection against lethal challenge with HSV-2 and induced a strong Th1-type immune response [109]. Skoberne et al. [110] prepared an experimental vaccine named GEN-003/MM-2, which contained gD2 and the IE protein ICP4/175K expressed in a baculovirus system. In combination with MM2 adjuvants (also called matrix M2 composed of cholesterol, phospholipids, and saponin), the vaccine showed immunotherapeutic effect in guinea pigs when it reduced the frequency of recurrent genital lesions upon HSV-2 challenge.

5.3. Viral vectored and DNA vaccines

The recent idea of DNA vaccines represents a progressive approach for immunization (reviewed at [111]). The advantage of DNA vaccines is that the immune response resembles to that following the administration of live-attenuated virus vaccines, but without the risk of reversion to viral phenotype. The viral DNA vaccines fall into two categories: viral DNA carriers and the recombinant plasmid vaccines (classical DNA vaccines). The viral DNA carriers are in fact nonpathogenic attenuated viruses that genome contains the inserted ORF fragments encoding the desired antigen. The best examples are either the recombinant adenovirus or vaccinia virus, which carry the HSV gD or gB genes. The latter glycoproteins become expressed in the immunized animal, that is, in the mice, which are then protected against HSV challenge [112115]. The recombinant vesicular stomatitis virus (VSV) was prepared from a plasmid containing the cDNA of the VSV RNA genome and the gD2 ORF US6. This recombinant virus induced a good cell-mediated immune response as well as anti-gD antibody formation. Immunized mice were protected against acute HSV-2 challenge, and the establishment of latency was also reduced [116]. Chiuppesi et al. [117] described the effect of immunization with feline lentiviral vector-based, herpes simplex virus 1 (HSV-1) glycoprotein B vaccine. This lentivirus construct induced HSV-1 antibody formation and also provided cross protection against lethal HSV-2 infection.

Many classical plasmid DNA vaccines were prepared with the inserted HSV DNA fragment. The majority of them encoded HSV-1 and/or HSV-2 glycoproteins such as gB, gD, gC, and gE; their efficacy was tested in mice, guinea pigs, and rabbits [90, 118124]. The results were obtained after immunization with plasmids encoding the glycoproteins gD and/or gB of either HSV-1 or HSV-2 alone or in combination. Especially the immunization of mice and/or guinea pigs with the gD-expressing plasmid protected against challenge with a lethal dose of HSV-2 [124128], but without clear-cut reduction of the latency rate. Better results were obtained with plasmids encoding both gB and gD [119, 129, 130]. The efficiency of DNA vaccines could be higher when adding cytokines such as IL-12 which induced the Th type 1 immune response [131, 132] similarly as the presence of IL-18 and/or RANTES [133, 134]. In contrast, the Th2-type response could be better achieved if the gD and IL-4-coding plasmid was used for immunization [133, 135]. The Th type 2 immune response is important for virus clearance from peripheral tissues including the infection of the eye, while the Th1-type response was involved in preventing the latency establishment [135]. The DNA vaccine-encoding gB when combined with DNA plasmid coding for cytokines IL-12 and IL-18 as adjuvant was efficient in prevention of the vaginal infection in mice: gB1/IL-12 and/or gB1/IL-18 elicited a local resistance of genital mucosa and protected mice against lethal HSV-2 challenge. The best results were observed with IL-12, while additional adjuvants did not enhance protection [136]. Another adjuvant tested with the DNA vaccines was the lipid adjuvant Vaxfectin®. In guinea pigs immunized with plasmid-expressing gD2 and VP11/VP12 as well as VP13/14 polypeptides along with a lipid adjuvant, a great prophylactic effect and the reduced HSV-2 replication in the genital tract of experimental animals was observed. The vaccine also showed immunotherapeutic properties when reducing the extent of latency (vDNA contents) in the dorsal root ganglia [137, 138]. Immunization with the recombinant plasmid encoding the fusion protein consisting of the gB and the CCL19 chemokine ORF induced both the Th type 1 and the Th type 2 responses including an increased local secretory IgA antibody production protecting mice against HSV-2 challenge [139]. The enhanced efficiency of DNA vaccines could be achieved by combining genes encoding the envelope glycoproteins with certain nonstructural HSV genes. The recombinant plasmid consisting of gD2, gB2, and ICP27 ORFs elicited a higher cellular as well as humoral response than the plasmid construct encoding the glycoproteins only. The construct in question also provided a higher protection against vaginal challenge with HSV-2 [140].

5.4. Live-attenuated HSV-1 and HSV-2 vaccines

Attenuated HSV vaccines are live HSV-1 and/or HSV-2 viruses derived from the wild-type strains by modifications of their virulence but keep the immunogenic properties. The best known deletion for HSV-1 was the removal of the UL22 ORF-encoding gH glycoprotein [141, 142] and/or the removal of the IR1 sequence encoding the protein γ34.5 gamma, a neurovirulence factor [143]. Furthermore, the deletion of the gE encoding by US8 gene, which is not needed for virus replication in cell culture but is inevitable for neural uptake [144, 145] along with the deletion of the UL41 gene, encoding the virion host shutoff (vhs) protein [146] might be an excellent solution. In this respect, the non-virulent HSZP strain with natural mutations altering the vhs protein, if deleted in the gE gene, would be of special advantage [147]. Another attempts to prepare a live-attenuated HSV strain were made by deleting nonstructural genes, such as the ORF UL54 (encodes the ICP27 IE polypeptide), the ORF RS1 (encodes the transactivation protein ICP4) [146, 148], and/or the ORF RL2 (encoding the ICP0 transactivation protein) as described by Halford et al. [149]. All the above-mentioned attenuated viruses was protective against HSV challenge in various animal models. The HSV-2 deleted in the gH gene designated as disabled infectious single cycle (DISC) and tested as therapeutic vaccine, but no convincing protection was found [150].

Several other attenuated HSV strains were prepared by deleting the nonstructural genes important for virulence. Deleted ORFs for such purpose were the following: (1) the UL23 ORF encoding the thymidine kinase (TK) as described by Morrison and Knipe [151], (2) the UL39 ORF encoding the large RR1 subunit [152], (3) the UL29 ORF encoding the ssDNA-binding ICP8 polypeptide [153], (4) the UL5 ORF encoding a protein of the primase/helicase complex [154, 155], and finally (5) the UL9 ORF encoding the ori-binding protein [156]. The TK minus recombinant R7017 was prepared from the w.t. strain F, which genome was, in addition, deleted in the IR1 ORF sequence encoding the γ34.5 and furthermore manipulated by inserting the gD2, gI2, and gG2 sequences along with a portion of the gE ORF (US4–US8). The TK-reversed recombinant virus was further modified by reinserting the TK gene (R7020 TK plus). Both recombinant viruses protected mice as well as guinea pigs against HSV-2 challenge [148]. The next UL39-/RR1-deleted virus was tested not only in animals but also in man [152, 157]. The phase I and phase II clinical trials showed partial protection against recurrences (in 37.5% of immunized individuals as compared with the placebo group) [157]. The attenuated strain deleted in the genes UL29 and UL5 (called dl29-5) protected mice against ocular infection with HSV-1 and guinea pigs against genital challenge with HSV-2 [158, 159]. Stanfield et al. [160] prepared an attenuated strain of HSV-2 called VC2, which had mutations in the gK gene and in the UL20 ORF (which is membrane protein inhibiting the neural uptake). This vaccine prevented genital infection upon vaginal challenge with HSV-2 and also inhibited latent infection in the lumbosacral dorsal root ganglia. Finally, the defective HSV-2 designated CJ9-gD2 was used to immunize mice. This virus had mutation in the UL9 ORF which encodes a vDNA replication protein and was deleted in the gD2 ORF (US9). The attenuated vaccine induced a higher antibody response in comparison with the gD2-alum/MPL subunit vaccine (used in a human trial) and protected mice against lethal challenge with HSV-2 [161].

6. Human vaccination trials: prophylactic and therapeutic HSV vaccines

Some of the vaccines mentioned above were tested not only in animal models but also in man at clinical trials. They were designed either as prophylactic vaccines, or they were destined for immunotherapeutic use, having been suggested for prevention or modulation of genital infection predominantly caused by HSV-2. The aim of prophylactic vaccine against HSV-2 infection is not only to prevent a clinical disease at primoinfection but also to interfere with the subsequent establishment of latency. The latter is a hard task, since the majority of vaccines just reduce the number of neurons which are getting HSV DNA carriers but does not fully prevent latent infection. Therefore, it is more reasonable to design a vaccine for immunotherapeutic use aimed to reduce the extent and the frequency of recurrent lesions.

The prophylactic vaccine should induce a satisfactory humoral as well as cell-mediated immune response. For such purpose, subunit, recombinant and DNA vaccines were tested. The first subunit vaccine tested was an HSV-2 glycoprotein mix (gB, gC, gG, gD, and gE) purified from infected chick embryo cells (this product is essentially similar to the subunit HSZP vaccine immuno mentioned above). As shown in 22 seronegative volunteers, this particular vaccine induced both the humoral and the cell-mediated specific response [162]. Alternatively, a similar vaccine (containing gB2 and gG2) was used to immunize 161 seronegative individuals with a less encouraging effect at phase II clinical trial [163]. The company Chiron Corporation sponsored the testing of a recombinant gB2/gG2 vaccine mixed with the MF59 adjuvant used for immunization of 137 persons. The efficiency of this vaccine was reported 9% only [53, 164]. Another study by GlaxoSmithKline uses a recombinant gD2 in combination with alum adjuvant and/or ASO4; in the latter trial, 7460 volunteers were selected as seronegative before the onset of the phase II trial. Immunization with the vaccine in question induced higher antibody titers than natural infection and also conferred partial protection against natural infection that had a milder course as compared to non-immunized controls [165]. At phase III trial, 847 seronegative individuals (no antibodies to HSV-1 as well as to HSV-2) and 1867 women showing antibodies to HSV-1 but not to HSV-2 were immunized with the same vaccine [166]. The results showed that 73–74% immunized women which were seronegative against both antigens have been protected against HSV-2 primoinfection (as detected by natural seroconversion), but the vaccine was not effective in man and in HSV-1 seropositive women. The GlaxoSmithKline company provided another trial, during which together 8323 seronegative women (for HSV-1 as well as HSV-2) were immunized. This follow-up was less encouraging when showing only 20% efficiency when measured by the seroconversion rate against natural genital HSV-2 infection and about 58% protection against HSV-1 genital infection [167]. In a multicenter study from 2013, the possible role of the abovementioned gD2-ASO4 vaccine in the etiology of stillbirth was evaluated. Together 19,727 pregnant women were immunized, from which 13.3% really had stillbirth in comparison with 11.00% of pregnant women in the control group that shows a clearly not significant effect [168].

The HSV DNA vaccine was also tested for prophylactic use. In the phase I trial, 62 seronegative women were immunized with the gD2 recombinant plasmid GENEVAX. T-cell-mediated response was found in one out of four volunteers [169]. Another prophylactic candidate vaccine tested was the HSV-2/HSV529-attenuated virus (deleted in genes UL5 and and UL29, also called dl5-29). This vaccine when tested in guinea pigs elicited a satisfactory humoral as well as T-cell response, conferred protection to HSV-2 challenge, and reduced the extent of latency in the regional lumbosacral ganglia [170]. The vaccine efficiency in human will be evaluated by the end of 2016.

As already mentioned, the therapeutic vaccines should decrease the frequency of recurrence episodes as well as their severity. To achieve this, the therapeutic vaccine must induce a potent and specific T-cell-mediated response. Several subunit vaccines were tested with an immunotherapeutic purpose, namely, the semipurified inactivated infected cell extracts, recombinant vaccines, and attenuated viruses. Probably, the first of the purified inactivated HSV-1-infected cell extract tested was the vaccine by Kutinová et al. [171]. A similar vaccine introduced by Skinner et al. [172] was used for immunization of 316 subjects with herpes disease who had recurrent blisters at least six times per year. Though this vaccine induced a detectable increase in the antibody levels and a satisfactory T-cell-mediated response, no difference in the frequency of recurrences was noted during the 1-year observation period. However, the vaccination accelerated the healing of the lesions, which had a less severe course in the immunized persons. Among the recombinant vaccines tested in a therapeutic context, the gD2 vaccine containing alum adjuvant should be mentioned first. This was used for immunization of 98 HSV-2 seropositive persons who had at least four but up to 14 recurrence episodes per year [173]. In this trial, the average of six recurrences per year in the placebo group was siginificantly higher than the average of four recurrences per year in the immunized group (p = 0.039). Furthermore, the gB2/gD2 vaccine mixed with the MF59 adjuvant was applied in another trial, in which 202 volunteers were immunized. At the phase II clinical trial, no difference was found in the titers of neutralizing antibodies during the observation period of 8 months, but there was a significant difference in the increased interval between the occurrences of the recurrent lesions following the immunization procedure [174]. Further trials were made using peptide vaccines. An overview of defined T-cell-based epitopes from HSV proteins was reported by Laing et al. ([175], Table 2).

Not all known T-cell based epitopes can induce protective immunity. Mapping of T cells in seropositive population found out that CD4+ T cells were mainly stimulated by tegument proteins UL21, UL46, UL47, UL49 and envelope glycoproteins gB and gD. It was observed that gB stimulates both CD4+ and CD8+ T cells, whereas gD induces stronger CD4+ T-cell-dependent immunity [58]. CD8+ T cells were stimulated by gD53–61, gD70–78, and gD278–286 peptides mainly in HLA-A*02:01-positive HSV-1 and HSV-2 seropositive healthy individuals [190]. Moreover, CD8+ T cells were also induced by tegument protein VP11/VP12 (gene UL46) that would account for another candidate protein to prepare effective HSV vaccine [61].

Protein (gene)HSV-1 T-cell epitopesHSV-2 T-cell epitopesB-cell epitopesReference
ICP0 (RL2)HumanHumanHuman[61]
VP5 (UL19)HumanHuman[176]
gH I (UL22)Mice[69]
TK (UL23)Human[177]
VP26 (UL25)Human, miceHuman[176, 178]
VP21 (UL26)HumanHuman[61]
gB (UL27)HumanHuman, miceHumanHumanMice[60, 61, 179181]
ICP8 (UL29)Human, miceHuman[61, 176]
ICP10 (UL39)Human, miceHumanHuman[61, 69, 176]
Vhs (UL41)Human, mice[61, 69]
gC (UL44)Human, miceMice[69, 180]
VP11/VP12 (UL46)HumanHuman, miceHuman, mice[61, 176, 182]
VP13/VP14 (UL47)HumanMiceHuman, mice[61, 176, 178]
VP16 (UL48)HumanHuman[61, 183, 184]
VP22 (UL49)HumanHuman, miceHuman, mice[61, 182, 183]
gK (UL53)MiceHuman, mice[61, 185, 186]
ICP27 (UL54)HumanHuman, mice[61, 187]
ICP22 (US1)Human[61]
gG (US4)MiceMice[69, 180]
gD (US6)Rabbit,
Rabbit, human,
Human, miceHuman, miceMice[161, 180, 188191]
gE (US8)HumanMice[178, 180]
ICP4 (RS1)HumanHumanHuman[61, 192, 193]

Table 2.

Overview of T- and B-cell HSV-1/HSV-2–specific epitopes as tested in human, mice, and rabbitsa


[i] - aModified according to Laing et al. [175].

Vaccine design also requires involvement of epitopes suitable for HLA binding by most people. In most ethnic groups, HLA-A*02:01 and HLA-B*07:02 belong to the most abundant HLA alleles [194]. HLA-A*02:01-restricted epitopes have been found in glycoprotein B (UL27 442–451), tegument protein VP13/VP14 (UL47 551–559), and tegument protein coded for UL25 (UL25 372–380) of HSV-2, and they stimulated CD8+ T-cell response [176, 195]. HLA-B*07:02-restricted epitopes have been reported for tegument protein VP22 (gene UL49) (HSV-2 UL49 49–57, HSV-2 UL49 82–90, HSV-2 UL49 99–108, HSV-2 UL49 131–140, and HSV-1 UL49 291–290) and stimulated CD8+ T-cell response [176]. Samandary et al. [196] found association with high prevalence of herpes infection and disease with the frequency of HLA-A*24, HLA-B*27, and HLA-B*58 alleles. In contrast, low prevalence of herpes infection and disease appeared associated with the high frequency of HLA-B*44 allele.

The effectivity of immune response is also depending on adjuvant type. Cooper et al. [189] found out that vaccine design and adjuvant type can have a significant effect on T-cell epitope utilization. Four epitopes within the gD2 molecule gD2 49–63, gD2 105–119, gD2 245–259 and gD2 333–347 were administered to mice with alum or IL-12. CD4+ T-cell response was induced in mice immunized with gD HSV-2 epitope gD2 245–259 and adjuvant alum. Mice immunized with IL-12 stimulate CD4+ T-cell response to HSV-2 epitope gD2 245–259 as well as to gD2 333–347 [189].

“Epitope” (peptide) vaccines that selectively stimulate T and B cells belong to other herpes simplex vaccine candidates. Wang et al. [180] prepared multi-epitope peptide vaccine that contained six B-cell epitopes from different glycoproteins of HSV-2 (gB2 466–473, gC2 216–223, gD2 6–18, gE2 483–491, gG2 572–579 and gI2 286–295), four CD4+ T-cell-based epitopes (gD2 21–28, gD2 205–224, gD2 245–259 and gB2 162–177) and two CD8+ T-cell-based epitopes (gD2 10–20 and gD2 268–276). All above-described epitopes were inserted into the extracellular fragment (1–290) of HSV-2 glycoprotein D to construct multi-epitope assembly peptides (MEAPs) by replacing some non-epitope amino acid sequences. The genes of the selected peptides were inserted into recombinant plasmid and expressed in E. coli strain BL21. The multi-epitope vaccine elicited in mice production of virus-neutralizing antibodies induced Th1 and Th2 immune response and protected mice against intravaginally induced lethal challenge of HSV-2 [180].

Many studies have focused on mapping of protective epitopes that stimulate immunity in asymptomatic individuals, that is, individuals without clinical findings of herpes infection. Analysis of IFN-γ-producing CD4+ T cells in HSV-1 seropositive individuals revealed that gB peptide epitopes (aa 166–180 and aa 666–680) were strongly recognized by CD4+ T cells from asymptomatic individuals, but not from symptomatic individuals. Inversely, CD4+ T cells from symptomatic individuals preferentially recognized gB (aa 661–675) [179]. Another study identified asymptomatic CD8+ T-cell epitopes from glycoprotein D (gD53–61, gD70–78 and gD278–286) [191]. It can be hypothesized that repertoire of T-cell-based epitopes determines either the development of HSV-1 (-2) clinical symptoms or asymptomatic viral shedding.

Finally, some investigators preferred the genetically attenuated virus vaccines, because they induce an immune response essentially similar to that following natural infection. For example, Casanova et al. [157] immunized the volunteers (32 persons having at least five recurrent lesions per year) with an attenuated HSV-2 virus deleted in the UL39 gene (the RR1 protein ORF). The phase I and phase II clinical trials showed the reduction of recurrent clinical manifestations by 37.5% as compared with the placebo-immunized group. Another clinical trial, which was performed in 2006, took the advantage of HSV-2-attenuated virus deleted in the gH gene (the so-called disabled infectious single cycle [DISC] virus). Unfortunately, no significant difference was found in the number of recurrences between the immunized and control mock-immunized subjects during the 1 year follow-up period [150]. The last phase I and II clinical trial which results will be announced nowadays concerns volunteers who had from two to nine recurrent lesions per year; they were immunized with a DNA vaccine-expressing gD2 in combination with the lipid adjuvant Vaxfectin [197].

7. Future perspectives of HSV-1/HSV-2 vaccination with the emphasis on the T-cell response

As described above, an effective vaccine against HSV-1 and HSV-2 infection that would prevent virus reactivation (therapeutic vaccine) should stimulate both humoral and cellular immunity mediated by CD4+ and CD8+ T cells. This immune response can be induced by live-attenuated virus, but such viruses are not safe because of their possible reversion back to the wild-type virus. Therefore, alternative approaches to develop effective herpes simplex vaccine have been attempted during the last decades. Nowadays, T-cell-inducing herpes simplex vaccines bearing “asymptomatic” immunodominant epitopes derived from HSV proteins were designed and tested [180]. Such vaccines possess many advantages over traditional vaccine like (1) induction of specific T-cell-based immunity, (2) inhibition of pathogenic immune response, and (3) safety.

Only few asymptomatic T-cell epitope-based vaccines were prepared and tested until now. The vaccine used by Wald et al. [198] contained 32 synthetic immunogenic HSV-2 peptides, linked to the heat shock protein Hsp 70 in combination with the QS-21 (contains a saponin) adjuvant. In 32 immunized volunteers, the vaccine elicited no complication, but induced a satisfactory CD4+/CD8+ T-lymphocyte response against a wide range of immunogenic HSV-2 peptides used in the stimulation tests in vitro. Chentoufi et al. [191] prepared CD8+ T-cell epitope-based vaccine containing three separate pairs of CD4–CD8 lipopeptides. Each of the lipopeptide contained one of the three asymptomatic immunodominant human CD8+ T-cell peptide epitopes from HSV-1 glycoprotein D (gD53–61, gD70–78, and gD278–286) that were joined with a human CD4+ T-cell peptide epitope (gD49–82). Humanized HLA-A*02:01 transgenic rabbits were immunized with a mixture of the three CD4–CD8 HSV-1 gD lipopeptides. Immunization induced an increased production of CD4+ and CD8+ T cells and protected rabbits against ocular HSV disease [191]. The same ASYMP vaccine was used for therapeutic vaccination of HLA transgenic rabbits infected by HSV-1. The vaccine induced production of HSV-specific CD8+ T cells that prevent HSV-1 reactivation ex vivo from latently infected explanted trigeminal ganglia. Moreover, the vaccine significantly reduced HSV-1 shedding and boosted the function of HSV-1 gD epitope-specific CD8+ T cells in draining lymph nodes, conjunctiva, and trigeminal ganglion [199].

The lipopeptide vaccines belong to another herpes simplex vaccine candidate. The lipopeptide vaccine can be easily produced and possesses some advantages over traditional vaccine such as safety and tolerance, bearing protective T-cell epitopes derived from HSV antigens, missing of non-immunogenic harmful epitopes, and missing of viral pathogenic proteins such as ICP47, and lipids have functioned as adjuvant. Previous studies observed that lipopeptide vaccine injected intranasally into mice induced mucosal and systemic B and Th1 immune response [191]. Other authors prefer attenuated genetically modified herpes simplex viruses as successfully vaccine candidates (Section 5.4).

8. Conclusions

In conclusion, the development of successfully therapeutical vaccine against HSV infection should respect the following recommendations: (1) the assessment of putative differences in the recognition of T- and/or B-cell epitopes from envelope, tegument, and regulatory HSV proteins in patients with recurrent herpes disease versus asymptomatic individuals; (2) the production of new safer adjuvants avoiding those claimed to cause the ASIA syndrome; (3) the induction of local mucosal immunity mediated by lipopeptides; (4) the use of humanized susceptible HLA transgenic mice as well as rabbits before human trials; and (5) at last but not least, the efficacy of human trials for an immunotherapeutic vaccine that should be made according to internationally accepted unified criteria in at least three groups of subjects (seronegative individuals, seropositive individuals without recurrent herpes disease, and volunteers with such disease). The tests should include the demonstration of elevated antibody titers after vaccination and in vitro testing of the T-lymphocyte response in a blastic transformation test as well as production of selected cytokines; 6. Finally, the application of the therapeutic vaccine in future human trials should be intracutaneous rather than by using other administration routes.


1 - Gruter W. Das Herpesvirus, seine aetiologische und klinische Bedeutung. Munch Med Wsch 1924, 71: 1058.
2 - Schneweis KE. Untersuchungen zur Typendifferenzierung des Herpesvirus hominis. Z Immun-Forsch 1962, 124: 24–28.
3 - Wildy P. Classification and nomenclature of viruses, pp. 33–34, In: Monographs in Virology, Melnick JL (ed), Karger, Basel, 1971.
4 - Lee EK, Coleman RM, Pereira L, Bailey PD, Tatsuno M, Nahmias AJ. Detection of herpes simplex virus type 2 specific antibody with glycoprotein G. J Clin Microbiol 1985, 22: 641–644.
5 - Liu F, Zhou ZH. Comparative virion structures of human herpesviruses, pp. 27–43, In: Arvin A, Campadelli-Fiume G, Mocarski E, Moore PS, Roizman B, Whitley R and Yamanishi K (eds). Human Herpesviruses: Biology, Therapy and Immunoprophylaxis. Cambridge University Press, 2007.
6 - Roizman B, Knipe DM, Whitley RJ. Herpes Simplex Viruses, pp. 1823–1897, In: Knipe DM, Howley PM (eds) Fields Virology, 6th ed., Walters Cluver/Lippincott Williams and Wilkins, Philadelphia, 2013.
7 - Knipe DM, Cliffe AR. Chromatin control of herpes simplex virus lytic and latent infection. Nat Rev Microbiol 2008, 6: 211–221. doi: 10.1038/nrmicro1794.
8 - Campadelli-Fiume G, Menotti L, Avitabile E, Gianni T. Viral and cellular contributions to herpes simplex virus entry into the cell. Curr Opin Virol 2012, 2: 28–36. doi: 10.1016/j.coviro.2011.12.001.
9 - Rajčáni J, Vojvodová A. The role of herpes simplex virus glycoproteins in the virus replication cycle. Acta Virol 1998, 42: 103–118.
10 - Gianni T, Amasio M, Campadelli-Fiume G. Herpes simplex virus gD forms distinct complexes with fusion executors gB and gH/gL in part through the C-terminal profusion domain. J Biol Chem 2009, 284: 17370–17382. doi: 10.1074/jbc.M109.005728.
11 - Satoh T, Arii J, Suenaga T, Wang J, Kogure A, Uehori J, Arase N, Shiratori I, Tanaka S, Kawaguchi Y, Spear PG, Lanier LL, Arase H. PILRα is a herpes simplex virus-1 entry co-receptor that associates with glycoprotein B. Cell 2008, 132: 935–944. doi: 10.1016/j.cell.2008.01.043.
12 - Stampfer SD, Lou H, Cohen GH, Eisenberg RJ, Heldwein EE. Structural basis of local, pH dependent conformational changes in glycoprotein B from herpes simplex virus type 1. J Virol 2010, 84: 12.924–12.933. doi: 10.1128/JVI.01750-10.
13 - Dingwell KS, Brunetti CR, Hendricks RL, Tang Q, Tang M, Rainbow AJ, Johnson DC. Herpes simplex virus glycoprotein E and I facilitate cell to cell spread in vivo and across junctions of cultured cells. J Virol 1994, 68: 834–845.
14 - Rajčáni J, Ďurmanová V. Early expression of herpes simplex virus (HSV) proteins and reactivation of latent infection. Folia Microbiol 2000, 45: 7–28
15 - Režuchová I, Kúdelová M, Ďurmanová V, Vojvodová A, Košovský J, Rajčáni J. Transcription at early stages of herpes simplex virus 1 infection and during reactivation. Intervirology 2003, 46: 25–34.
16 - Stevens JG, Wagner EK, Devi-Rao GB, Cook ML, Feldman LT. RNA complementary to a herpesvirus alpha gene mRNA is prominent in latently infected neurons. Science 1987, 235: 1056–1059.
17 - Ahmed MM, Lock C, Millerm G. Regions of the herpes simplex virus type 1 latency-associated transcript that protect cells from apoptosis in vitro and protect neuronal cells in vivo. J Virol 2002, 76: 717–722.
18 - Perng GC, Jones C, Ciacci-Zanella J, Stone M, Henderson G, Yukht A, Slanina SM, Hofman FM, Ghiasi H, Nesburn AB, Wechsler SL. Virus induced neuronal apoptosis blocked by the herpes simplex virus latency associated transcript (LAT). Science 2000, 287: 1500–1503.
19 - Perng GC, Maguen B, Jin L, Mott KR, Osorio N, Slanina SM, Yukht A, Ghiasi H, Nesburn AB, Inman M, Henderson G, Jones C, Wechsler SL. A gene capable of blocking apoptosis can substitute for herpes simplex virus type 1 latency associated transcript gene and restore wild type reactivation levels. J Virol 2002, 76: 1224–1235
20 - Efstathiou S, Minson AC, Field HJ, Anderson JR, Wildy P. Detection of herpes simplex virus specific DNA sequences in latently infected mice and in humans. J Virol 1986, 32: 446–455.
21 - Kristie TM and Roizman B. Differentiation and DNA contact points to the host proteins binding at the cis site of for virion-mediated induction of herpes simplex virus 1 alpha genes. J Virol 1988, 62: 1145–1157
22 - Lillycrop KA, Budrahan VS, Lakin ND, Terrenghi G, Wood JN, Polak JM, Latchman DS. A novel POU family transcription factor is closely related to Brn-3 but has a distinct expression pattern in neuronal cells. Nucleic Acids Res 1992, 20: 5093–5096.
23 - Lillycrop KA, Estridge JK, Latchman DS. Functional interaction between different isoforms of the Oct-2 transcription factor expressed in neuronal cells. Biochem J 1994, 298: 245–248.
24 - Preston CM, Efstathiou S. Molecular basis of HSV latency and reactivation, pp. 602–615, In: Arvin A, Campadelli-Fiume G, Mocarski E, Moore PS, Roizman B, Whitley R, Yamanishi K. (eds.) Human Herpesviruses, Biology, Therapy and Immunoprophylaxis, Cambridge University Press, 2007.
25 - Fries LF, Friedman HM, Cohen GH, Eisenberg RJ, Hammer CH, Frank MM. Glycoprotein C of herpes simplex virus 1 is an inhibitor of the complement cascade. J Immunol 1986, 137: 1636–1641.
26 - Hill TJ, Field HJ, Blyth WA. Acute and recurrent infection with herpes simplex virus in the mouse: a model for studying latency and recurrent disease. J Gen Virol 1975, 28: 341–353.
27 - Hill TJ, Blyth HJ. An alternative theory of herpes simplex recurrence and a possible role for prostaglandins. Lancet 1976, 1: 397–398.
28 - Walz M, Price R, Notkins A. Latent ganglionic infection with herpes simplex virus types 1 and 2: viral reactivation after neurectomy. Science 1974, 184: 1185–1187.
29 - Kúdelová M, Rajčáni J. Herpes simplex virus and human CNS infection, pp. 169–214, In: Singh SK, Ružek D. (eds), Neuroviral Infection, 1st ed., Taylor and Francis Group CRC, 2013.
30 - Whitley R, Kimberlin D, Prober Ch. HSV 1 and 2: pathogenesis and disease. pp. 569–601, In: Arvin A, Campadelli-Fiume G, Mocarski E, Moore PS, Roizman B, Whitley R, Yamanishi K. (eds.). Human Herpesviruses, Biology, Therapy and Immunoprophylaxis, Cambridge University Press, 2007.
31 - Svensson A, Bellner L, Magnusson M, Eriksson K. Role of IFN-alpha/beta signaling in the prevention of genital herpes virus type 2 infection. J Reprod Immunol 2007, 74: 114–123.
32 - Conrady CD, Halford WP, Carr DJ. Loss of the type I interferon pathway increases vulnerability of mice to genital herpes simplex virus 2 infection. J Virol 2011, 85: 1625–1633. doi: 10.1128/JVI.01715-10.
33 - Kim M, Osborne NR, Zeng W, Donaghy H, McKinnon K, Jackson DC, Cunningham AL. Herpes simplex virus antigens directly activate NK cells via TLR2, thus facilitating their presentation to CD4 T lymphocytes. J Immunol 2012, 188: 4158–4170. doi: 10.4049/jimmunol.1103450.
34 - Lundberg P, Welander P, Han X, Cantin E. Herpes simplex virus type 1 DNA is immunostimulatory in vitro and in vivo. J Virol 2003, 77: 11158–11169.
35 - Guo Y, Audry M, Ciancanelli M, Alsina L, Azevedo J, Herman M, Anguiano E, Sancho-Shimizu V, Lorenzo L, Pauwels E, Philippe PB, Pérez de Diego R, Cardon A, Vogt G, Picard C, Andrianirina ZZ, Rozenberg F, Lebon P, Plancoulaine S, Tardieu M, Valérie D, Jouanguy E, Chaussabel D, Geissmann F, Abel L, Casanova JL, Zhang SY. Herpes simplex virus encephalitis in a patient with complete TLR3 deficiency: TLR3 is otherwise redundant in protective immunity. J Exp Med 2011, 208: 2083–2098. doi: 10.1084/jem.20101568.
36 - Herman M, Ciancanelli M, Ou YH, Lorenzo L, Klaudel-Dreszler M, Pauwels E, Sancho-Shimizu V, Pérez de Diego R, Abhyankar A, Israelsson E, Guo Y, Cardon A, Rozenberg F, Lebon P, Tardieu M, Heropolitanska-Pliszka E, Chaussabel D, White MA, Abel L, Zhang SY, Casanova JL. Heterozygous TBK1 mutations impair TLR3 immunity and underlie herpes simplex encephalitis of childhood. J Exp Med 2012, 209: 1567–1582. doi: 10.1084/jem.20111316.
37 - Ank N, West H, Bartholdy C, Eriksson K, Thomsen AR, Paludan SR. Lambda interferon (IFN-lambda), a type III IFN, is induced by viruses and IFNs and displays potent antiviral activity against select virus infections in vivo. J Virol 2006, 80: 4501–4509.
38 - Li J, Hu S, Zhou L, Ye L, Wang X, Ho J, Ho W. Interferon lambda inhibits herpes simplex virus type I infection of human astrocytes and neurons. Glia 2011, 59: 58–67. doi: 10.1002/glia.21076.
39 - Lopušná K, Režuchová I, Kabát P, Kúdelová M. Interferon lambda induces antiviral response to herpes simplex virus 1 infection. Acta Virol 2014, 58: 325–332.
40 - Grubor-Bauk B, Arthur JL, Mayrhofer G. Importance of NKT cells in resistance to herpes simplex virus, fate of virus-infected neurons, and level of latency in mice. J Virol 2008, 82: 11073–11083. doi: 10.1128/JVI.00205-08.
41 - Adler H, Beland JL, Del-Pan NC, Kobzik L, Sobel RA, Rimm IJ. In the absence of T cells, natural killer cells protect from mortality due to HSV-1 encephalitis. J Neuroimmunol 1999, 93: 208–213.
42 - Milligan GN. Neutrophils aid in protection of the vaginal mucosae of immune mice against challenge with herpes simplex virus type 2. J Virol 1999, 73: 6380–6386.
43 - Kodukula P, Liu T, Rooijen NV, Jager MJ, Hendricks RL. Macrophage control of herpes simplex virus type 1 replication in the peripheral nervous system. J Virol 1999, 162: 2895–2905.
44 - Buc M. Interferon gamma (IFN-gamma), p. 95, In: Basic and Clinical Immunology, Comenius University, Bratislava, 2008.
45 - Kassim SH, Rajasagi NK, Zhao X, Chervenak R, Jennings SR. In vivo ablation of CD11c-positive dendritic cells increases susceptibility to herpes simplex virus type 1 infection and diminishes NK and T-cell responses. J Virol 2006, 80: 3985–3993.
46 - Hochrein H, Schlatter B, O'Keeffe M, Wagner C, Schmitz F, Schiemann M, Bauer S, Suter M, Wagner H. Herpes simplex virus type-1 induces IFN-alpha production via Toll-like receptor 9-dependent and -independent pathways. Proc Natl Acad Sci U S A 2004, 101: 11416–11421.
47 - den Haan JM, Bevan MJ. Antigen presentation to CD8+ T cells: cross-priming in infectious diseases. Curr Opin Immunol 2001, 13: 437–441.
48 - Ramirez MC, Sigal LJ. The multiple routes of MHC-I cross-presentation. Trends Microbiol 2004, 12: 204–207.
49 - Lubinski J, Wang L, Mastellos D, Sahu A, Lambris JD, Friedman HM. In vivo role of complement-interacting domains of herpes simplex virus type 1 glycoprotein gC. J Exp Med 1999, 190: 1637–1646.
50 - McDermott MR, Brais LJ, Evelegh MJ. Mucosal and systemic antiviral antibodies in mice inoculated intravaginally with herpes simplex virus type 2. J Gen Virol 1990, 71: 1497–1504.
51 - Dudley KL, Bourne N, Milligan GN. Immune protection against HSV-2 in B-cell-deficient mice. Virology 2000, 270: 454–463.
52 - Morrison LA, Zhu L, Thebeau LG. Vaccine-induced serum immunoglobin contributes to protection from herpes simplex virus type 2 genital infection in the presence of immune T cells. J Virol 2001, 75: 1195–1204.
53 - Corey L, Langenberg AG, Ashley R, Sekulovich RE, Izu AE, Douglas JM Jr., Handsfield HH, Warren T, Marr L, Tyring S, DiCarlo R, Adimora AA, Leone P, Dekker CL, Burke RL, Leong WP, Straus SE. Recombinant glycoprotein vaccine for the prevention of genital HSV-2 infection: two randomized controlled trials. Chiron HSV Vaccine Study Group. JAMA 1999, 282: 331–340.
54 - Handunnetthi L, Ramagopalan SV, Ebers GC, Knight JC. Regulation of major histocompatibility complex class II gene expression, genetic variation and disease. Genes Immun 2010, 11: 99–112. doi: 10.1038/gene.2009.83.
55 - Milligan GN, Bernstein DI. Interferon-gamma enhances resolution of herpes simplex virus type 2 infection of the murine genital tract. Virology 1997, 229: 259–268.
56 - Milligan GN, Bernstein DI, Bourne N. T lymphocytes are required for protection of the vaginal mucosae and sensory ganglia of immune mice against reinfection with herpes simplex virus type 2. J Immunol 1998, 160: 6093–6100.
57 - Gill N, Ashkar AA. Overexpression of interleukin-15 compromises CD4-dependent adaptive immune responses against herpes simplex virus 2. J Virol 2009, 83: 918–926. doi: 10.1128/JVI.01282-08.
58 - Koelle DM, Schomogyi M, McClurkan C, Reymond SN, Chen HB. CD4 T-cell responses to herpes simplex virus type 2 major capsid protein VP5: comparison with responses to tegument and envelope glycoproteins. J Virol 2000, 74: 11422–11425.
59 - Zhu J, Koelle DM, Cao J, Vazquez J, Huang ML, Hladik F, Wald A, Corey L. Virus-specific CD8+ T cells accumulate near sensory nerve endings in genital skin during subclinical HSV-2 reactivation. J Exp Med 2007, 204: 595–603.
60 - Khanna KM, Bonneau RH, Kinchington PR, Hendricks RL. Herpes simplex virus-specific memory CD8+ T cells are selectively activated and retained in latently infected sensory ganglia. Immunity 2003, 18: 593–603.
61 - Jing L, Haas J, Chong TM, Bruckner JJ, Dann GC, Dong L, Marshak JO, McClurkan CL, Yamamoto TN, Bailer SM, Laing KJ, Wald A, Verjans GM, Koelle DM. Cross-presentation and genome-wide screening reveal candidate T cells antigens for a herpes simplex virus type 1 vaccine. J Clin Invest 2012, 122: 654–763. doi: 10.1172/JCI60556.
62 - Gebhardt BM, Hill JM. Cellular neuroimmunologic responses to ocular herpes simplex virus infection. J Neuroimmunol 1990, 28: 227–236.
63 - Liu T, Tang Q, Hendricks RL Inflammatory infiltration of the trigeminal ganglion after herpes simplex virus type 1 corneal infection. J Virol 1996, 70: 264–271 (1996).
64 - Liu T, Khanna KM, Chen X, Fink DJ, Hendricks RL. CD8(+) T cells can block herpes simplex virus type 1 (HSV-1) reactivation from latency in sensory neurons. J Exp Med 2000, 191: 1459–1466.
65 - Theil D, Derfuss T, Paripovic I, Herberger S, Meinl E, Schueler O, Strupp M, Arbusow V, Brandt T. Latent herpesvirus infection in human trigeminal ganglia causes chronic immune response. Am J Pathol 2003, 163: 2179–2184.
66 - Knickelbein JE, Khanna KM, Yee MB, Baty CJ, Kinchington PR, Hendricks RL. Noncytotoxic lytic granule-mediated CD8+ T cell inhibition of HSV-1 reactivation from neuronal latency. Science 2008, 322: 268–271. doi: 10.1126/science.1164164.
67 - Decman V, Kinchington PR, Harvey SA, Hendricks RL. Gamma interferon can block herpes simplex virus type 1 reactivation from latency, even in the presence of late gene expression. J Virol 2005, 79: 10339–10347.
68 - Mueller SN, Heath W, McLain JD, Carbone FR, Jones CM. Characterization of two TCR transgenic mouse lines specific for herpes simplex virus. Immunol Cell Biol 2002, 80: 156–163.
69 - St Leger AJ, Peters B, Sidney J, Sette A, Hendricks RL. Defining the herpes simplex virus-specific CD8+ T cell repertoire in C57BL/6 mice. J Immunol 2011, 186: 3927–3933. doi: 10.4049/jimmunol.1003735.
70 - Derfuss T, Segerer S, Herberger S, Sinicina I, Hüfner K, Ebelt K, Knaus HG, Steiner I, Meinl E, Dornmair K, Arbusow V, Strupp M, Brandt T, Theil D. Presence of HSV-1 immediate early genes and clonally expanded T-cells with a memory effector phenotype in human trigeminal ganglia. Brain Pathol 2007, 17: 389–398.
71 - van Velzen M, Jing L, Osterhaus AD, Sette A, Koelle DM, Verjans GM. Local CD4 and CD8 T-cell reactivity to HSV-1 antigens documents broad viral protein expression and immune competence in latently infected human trigeminal ganglia. PLoS Pathog 2013, 9: e1003547. doi: 10.1371/journal.ppat.1003547.
72 - Egan KP, Wu S, Wigdahl B, Jennings SR. Immunological control of herpes simplex virus infections. J Neurovirol 2013, 19: 328–345. doi: 10.1007/s13365-013-0189-3.
73 - Kern A, Schiff B. Vaccine therapy in recurrent herpes simplex. Arch Dermatol 1964, 89: 844–845.
74 - Koelle D. HSV 1 and 2 immunobiology and host response, pp. 616–641, In: Arvin A, Campadelli-Fiume G, Mocarski E, Moore PS, Roizman B, Whitley R, Yamanishi K (eds). Human Herpesviruses: Biology, Therapy and Immunoprophylaxis. Cambridge University Press, 2007.
75 - Skinner GR, Buchan A, Hartley CE, Turner SP, Williams DR. The preparation, efficacy and safety of 'antigenoid' vaccine NFU1 (S-L+) MRC toward prevention of herpes simplex virus infections in human subjects. Med Microbiol Immunol 1980, 169: 39–51.
76 - Dundarov S, Andonov P, Bakalov B, Nechev K, Tomov C. Immunotherapy with inactivated polyvalent herpes vaccines. Dev Biol Stand 1982, 52: 351–358.
77 - Scriba M. Animal studies on the efficacy of vaccination against recurrent herpes. Med Microbiol Immunol 1982, 171: 33–40.
78 - Kutinová V, Vonka V, Řezáčová D. Production and some properties of antigen extracts for neutralizing of herpes simplex virus. Acta Virol 1977, 19: 189–197.
79 - Rajčáni J, Kutinová L, Vonka V. Restriction of latent herpes virus infection of rabbits immunized with subviral herpes simplex virus vaccine. Acta Virol 1980, 24: 183–193.
80 - Morello CS, Kraynyak KA, Levinson MS, Chen Z, Lee KF, Spector DH. Inactivated HSV-2 in MPL/alum adjuvant provides nearly complete protection against genital infection and shedding following long term challenge and rechallenge. Vaccine 2012, 30: 6541–6550. doi: 10.1016/j.vaccine.2012.08.049.
81 - Dundarov S, Andonov P. Seventeen years of application of herpes vaccines in Bulgaria. Acta Virol 1994, 38: 205–208.
82 - Hilfenhaus J, Christ H, Köhler R, Moser H, Kirchner H, Levy HB. Protectivity of herpes simplex virus antigens: studies in mice on the adjuvant effect of PICLC and on the dependence of protection on T cell competence. Med Microbiol Immunol 1981, 169: 225–235.
83 - Thomson TA, Hilfenhaus J, Moser H, Morahan PS. Comparison of effects of adjuvants on efficacy of virion envelope herpes simplex virus vaccine against labial infection of BALB/c mice. Infect Immun 1983, 41: 556–562.
84 - Metcalf JF, Whitley RJ. Protective immunity against herpetic ocular disease in an outbred mouse model. Curr Eye Res 1987, 6: 167–171.
85 - Rajčáni J, Sabó A, Mucha V, Košťál M, Compel P. Herpes simplex virus type 1 envelope subunit vaccine not only protects against lethal virus challenge, but also may restrict latency and virus reactivation. Acta Virol 1995, 39: 37–49.
86 - Myers MG, Bernstein DI, Harrison CJ, Stanberry LR. Herpes simplex virus glycoprotein treatment of recurrent genital herpes reduces cervicovaginal virus shedding in guinea pigs. Antiviral Res 1988, 10: 83–88.
87 - Blacklaws BA, Krishna S, Minson AC, Nash AA. Immunogenicity of herpes simplex virus type 1 glycoproteins expressed in vaccinia virus recombinants. Virology 1990, 177: 727–736.
88 - York LJ, Giorgio DP, Mishkin EM. Immunomodulatory effects of HSV2 glycoprotein D in HSV1 infected mice: implications for immunotherapy of recurrent HSV infection. Vaccine 1995, 13: 1706–1712.
89 - Mosko T, Kosovský J, Rezuchová I, Durmanová V, Kúdelová M, Rajcáni J. Expression of herpes simplex virus 1 glycoprotein D in prokaryotic and eukaryotic cells. Acta Virol 2004, 48: 97–107.
90 - Durmanová V, Mosko T, Sapák M, Kosovský J, Rezuchová I, Buc M, Rajcáni J. Efficacy of recombinant herpes simplex virus 1 glycoprotein D candidate vaccines in mice. Acta Microbiol Immunol Hung 2006, 53: 459–477.
91 - Long D, Madara TJ, Ponce de Leon M, Cohen GH, Montgomery PC, Eisenberg RJ. Glycoprotein D protects mice against lethal challenge with herpes simplex virus types 1 and 2. Infect Immun 1984, 43: 761–764.
92 - Roberts PL, Duncan BE, Raybould TJ, Watson DH. Purification of herpes simplex virus glycoproteins B and C using monoclonal antibodies and their ability to protect mice against lethal challenge. J Gen Virol 1985, 66: 1073–1085.
93 - Kino Y, Eto T, Nishiyama K, Ohtomo N, Mori R. Immunogenicity of purified glycoprotein gB of herpes simplex virus. Arch Virol 1986, 89: 69–80.
94 - Manservigi R, Grossi MP, Gualandri R, Balboni PG, Marchini A, Rotola A, Rimessi P, Di Luca D, Cassai E, Barbanti-Brodano G. Protection from herpes simplex virus type 1 lethal and latent infections by secreted recombinant glycoprotein B constitutively expressed in human cells with a BK virus episomal vector. J Virol 1990, 64: 431–436.
95 - Broker M, Abel KJ, Köhler R, Hilfenhaus J, Amann E. Escherichia coli-derived envelope protein gD but not gC antigens of herpes simplex virus protect mice against a lethal challenge with HSV-1 and HSV-2. Med Microbiol Immunol 1990, 179: 145–159.
96 - Byars NE, Fraser-Smith EB, Pecyk RA, Welch M, Nakano G, Burke RL, Hayward AR, Allison AC. Vaccinating guinea pigs with recombinant glycoprotein D of herpes simplex virus in an efficacious adjuvant formulation elicits protection against vaginal infection. Vaccine 1994, 12: 200–209.
97 - O'Hagan D, Goldbeck C, Ugozzoli M, Ott G, Burke RL. Intranasal immunization with recombinant gD2 reduces disease severity and mortality following genital challenge with herpes simplex virus type 2 in guinea pigs. Vaccine 1999, 17: 2229–2236.
98 - Bourne N, Bravo FJ, Francotte M, Bernstein DI, Myers MG, Slaoui M, Stanberry LR. Herpes simplex virus (HSV) type 2 glycoprotein D subunit vaccines and protection against genital HSV-1 or HSV-2 disease in guinea pigs. J Infect Dis 2003, 187: 542–549.
99 - Manservigi R, Boero A, Argnani R, Caselli E, Zucchini S, Miriagou V, Mavromara P, Cilli M, Grossi MP, Balboni PG, Cassai E. Immunotherapeutic activity of a recombinant combined gB-gD-gE vaccine against recurrent HSV-2 infections in a guinea pig model. Vaccine 2005, 23: 865–872.
100 - Durmanová V, Sapák M, Kosovský J, Rezuchová I, Kúdelová M, Buc M, Rajcáni J. Immune response and cytokine production following immunization with experimental herpes simplex virus 1 (HSV-1) vaccines. Folia Microbiol (Praha) 2008, 53: 73–83. doi: 10.1007/s12223-008-0011-4.
101 - Fotouhi F, Soleimanjahi H, Roostaee MH, Dalimi Asl A. Expression of the herpes simplex virus type 2 glycoprotein D in baculovirus expression system and evaluation of its immunogenicity in guinea pigs. Iran Biomed J 2008, 12: 59–66.
102 - Awasthi S, Balliet JW, Flynn JA, Lubinski JM, Shaw CE, DiStefano DJ, Cai M, Brown M, Smith JF, Kowalski R, Swoyer R, Galli J, Copeland V, Rios S, Davidson RC, Salnikova M, Kingsley S, Bryan J, Casimiro DR, Friedman HM. Protection provided by a herpes simplex virus 2 (HSV-2) glycoprotein C and D subunit antigen vaccine against genital HSV-2 infection in HSV-1-seropositive guinea pigs. J Virol 2014, 88: 2000–2010. doi: 10.1128/JVI.03163-13.
103 - Awasthi S, Mahairas GG, Shaw CE, Huang ML, Koelle DM, Posavad C, Corey L, Friedman HM. A dual-modality herpes simplex virus 2 vaccine for preventing genital herpes by using glycoprotein C and D subunit antigens to induce potent antibody responses and adenovirus vectors containing capsid and tegument proteins as T cell immunogens. J Virol 2015, 89: 8497–8509. doi: 10.1128/JVI.01089-15.
104 - Inoue T, Inoue Y, Nakamura T, Yoshida A, Inoue Y, Tano Y, Shimomura Y, Fujisawa Y, Aono A, Hayashi K. The effect of immunization with herpes simplex virus glycoprotein D fused with interluekin-2 against murine herpetic keratitis. Nihon Ganka Gakkai Zasshi 2001, 105: 223–229.
105 - Mohamedi SA, Heath AW, Jennings R. A comparison of oral and parenteral routes for therapeutic vaccination with HSV-2 ISCOMs in mice; cytokine profiles, antibody responses and protection. Antiviral Res 2001, 49: 83–99.
106 - Burke RL, Goldbeck C, Ng P, Stanberry L, Ott G, Van Nest G. The influence of adjuvant on the therapeutic efficacy of a recombinant genital herpes vaccine. J Infect Dis 1994, 170: 1110–1119.
107 - Shoenfeld Y, Agmont-Levine N. ASIA: autoimmune/inflammatory syndrome induced by adjuvants. J Autoimmun 2011, 36: 4–8. doi: 10.1016/j.jaut.2010.07.003.
108 - Lavelle EC, Grant G, Pusztai A, Pfüller U, Leavy O, McNeela E, Mills KH, O'Hagan DT. Mistletoe lectins enhance immune responses to intranasally co-administered herpes simplex virus glycoprotein D2. Immunology 2002, 107: 268–274.
109 - Cortesi R, Ravani L, Rinaldi F, Marconi P, Drechsler M, Manservigi M, Argnani R, Menegatti E, Esposito E, Manservigi R. Intranasal immunization in mice with non-ionic surfactants vesicles containing HSV immunogens: a preliminary study as possible vaccine against genital herpes. Int J Pharm 2013, 440: 229–237. doi: 10.1016/j.ijpharm.2012.06.042.
110 - Skoberne M, Cardin R, Lee A, Kazimirova A, Zielinski V, Garvie D, Lundberg A, Larson S, Bravo FJ, Bernstein DI, Flechtner JB, Long D. An adjuvanted herpes simplex virus 2 subunit vaccine elicits a T cell response in mice and is an effective therapeutic vaccine in guinea pigs. J Virol 2013, 87: 3930–3942. doi: 10.1128/JVI.02745-12.
111 - Rajčáni J, MoškoT, Režuchová, I. Current developments in viral DNA vaccines: shall they solve the unsolved? Rev Med Microbiol 2005, 15, 1–23.
112 - Cantin EM, Eberle R, Baldick JL, Moss B, Willey DE, Notkins AL, Openshaw H. Expression of herpes simplex virus 1 glycoprotein B by a recombinant vaccinia virus and protection of mice against lethal herpes simplex virus 1 infection. Proc Natl Acad Sci U S A 1987, 84: 5908–5912.
113 - Willey DE, Cantin EM, Hill LR, Moss B, Notkins AL, Openshaw H. Herpes simplex virus type 1-vaccinia virus recombinant expressing glycoprotein B: protection from acute and latent infection. J Infect Dis 1988, 158: 1382–1386.
114 - Wachsman M, Aurelian L, Smith CC, Perkus ME, Paoletti E. Regulation of expression of herpes simplex virus (HSV) glycoprotein D in vaccinia recombinants affects their ability to protect from cutaneous HSV-2 disease. J Infect Dis 1989, 159: 625–634.
115 - Bernstein DI. Effect of route of vaccination with vaccinia virus expressing HSV-2 glycoprotein D on protection from genital HSV-2 infection. Vaccine 2000, 18: 1351–1358.
116 - Natuk RJ, Cooper D, Guo M, Calderon P, Wright KJ, Nasar F, Witko S, Pawlyk D, Lee M, DeStefano J, Tummolo D, Abramovitz AS, Gangolli S, Kalyan N, Clarke DK, Hendry RM, Eldridge JH, Udem SA, Kowalski J. Recombinant vesicular stomatitis virus vectors expressing herpes simplex virus type 2 gD elicit robust CD4+ Th1 immune responses and are protective in mouse and guinea pig models of vaginal challenge. J Virol 2006, 80: 4447–4457.
117 - Chiuppesi F, Vannucci L, De Luca A, Lai M, Matteoli B, Freer G, Manservigi R, Ceccherini-Nelli L, Maggi F, Bendinelli M, Pistello M. A lentiviral vector-based, herpes simplex virus 1 (HSV-1) glycoprotein B vaccine affords cross-protection against HSV-1 and HSV-2 genital infections. J Virol 2012, 86: 6563–6574. doi: 10.1128/JVI.00302-12.
118 - Daheshia M, Kuklin N, Manickan E, Chun S, Rouse BT. Immune induction and modulation by topical ocular administration of plasmid DNA encoding antigens and cytokines. Vaccine 1998, 16: 1103–1110.
119 - Nass PH, Elkins KL, Weir JP. Antibody response and protective capacity of plasmid vaccines expressing three different herpes simplex virus glycoproteins. J Infect Dis 1998, 178: 611–617.
120 - Stanberry LR, Cunningham AL, Mindel A, Scott LL, Spruance SL, Aoki FY, Lacey CJ. Prospects for control of herpes simplex virus disease through immunization. Clin Infect Dis 2000, 30: 549–566.
121 - Baghian A, Chouljenko VN, Dauvergne O, Newmant MJ, Baghian S, Kousoulas KG. Protective immunity against lethal HSV-1 challenge in mice by nucleic acid-based immunisation with herpes simplex virus type-1 genes specifying glycoproteins gB and gD. J Med Microbiol 2002, 51: 350–357.
122 - Osorio Y, Cohen J, Ghiasi H. Improved protection from primary ocular HSV-1 infection and establishment of latency using multigenic DNA vaccines. Invest Ophthalmol Vis Sci 2004, 45: 506–514
123 - Hoshino Y, Dalai SK, Wang K, Pesnicak L, Lau TY, Knipe DM, Cohen JI, Straus SE. Comparative efficacy and immunogenicity of replication-defective, recombinant glycoprotein, and DNA vaccines for herpes simplex virus 2 infections in mice and guinea pigs. J Virol 2005, 79: 410–418.
124 - Ghaemi A, Soleimanjahi H, Bamdad T, Soudi S, Arefeian E, Hashemi SM, Ebtekar M. Induction of humoral and cellular immunity against latent HSV-1 infections by DNA immunization in BALB/c mice. Comp Immunol Microbiol Infect Dis 2007, 30: 197–210.
125 - Bourne N, Milligan GN, Schleiss MR, Bernstein DI, Stanberry LR. DNA immunization confers protective immunity on mice challenged intravaginally with herpes simplex virus type 2. Vaccine 1996, 14: 1230–1234.
126 - McClements WL, Armstrong ME, Keys RD, Liu MA. The prophylactic effect of immunization with DNA encoding herpes simplex virus glycoproteins on HSV-induced disease in guinea pigs. Vaccine 1997, 15: 857–860.
127 - Strasser JE, Arnold RL, Pachuk C, Higgins TJ, Bernstein DI. Herpes simplex virus DNA vaccine efficacy: effect of glycoprotein D plasmid constructs. J Infect Dis 2000, 182: 1304–1310.
128 - Jazayeri M, Soleimanjahi H, Fotouhi F, Pakravan N. Comparison of intramuscular and footpad subcutaneous immunization with DNA vaccine encoding HSV-gD2 in mice. Comp Immunol Microbiol Infect Dis 2009, 32: 453–461. doi: 10.1016/j.cimid.2008.05.002.
129 - Lee HH, Cha SC, Jang DJ, Lee JK, Choo DW, Kim YS, Uh HS, Kim SY. Immunization with combined HSV-2 glycoproteins B2: D2 gene DNAs: protection against lethal intravaginal challenges in mice. Virus Genes 2002, 25: 179–188.
130 - Domingo C, Gadea I, Pardeiro M, Castilla C, Fernández S, Fernández-Clua MA, De la Cruz Troca JJ, Punzón C, Soriano F, Fresno M, Tabarés E. Immunological properties of a DNA plasmid encoding a chimeric protein of herpes simplex virus type 2 glycoprotein B and glycoprotein D. Vaccine 2003, 21: 3565–3574
131 - Sin JI, Kim JJ, Arnold RL, Shroff KE, McCallus D, Pachuk C, McElhiney SP, Wolf MW, Pompa-de Bruin SJ, Higgins TJ, Ciccarelli RB, Weiner DB. IL-12 gene as a DNA vaccine adjuvant in a herpes mouse model: IL-12 enhances Th1-type CD4+ T cell-mediated protective immunity against herpes simplex virus-2 challenge. J Immun 1999, 162: 2912–2921.
132 - Cooper D, Pride MW, Guo M, Cutler M, Mester JC, Nasar F, She J, Souza V, York L, Mishkin E, Eldridge J, Natuk RJ. Interleukin-12 redirects murine immune responses to soluble or aluminum phosphate adsorbed HSV-2 glycoprotein D towards Th1 and CD4+ CTL responses. Vaccine 2004, 23: 236–246.
133 - Sin JI, Kim JJ, Boyer JD, Ciccarelli RB, Higgins TJ, Weiner DB. In vivo modulation of vaccine-induced immune responses toward a Th1 phenotype increases potency and vaccine effectiveness in a herpes simplex virus type 2 mouse model. J Virol 1999, 73: 501–509.
134 - Zhu M, Xu X, Liu H, Liu X, Wang S, Dong F, Yang B, Song G. Enhancement of DNA vaccine potency against herpes simplex virus 1 by co-administration of an interleukin-18 expression plasmid as a genetic adjuvant. J Med Microbiol 2003, 52: 223–228.
135 - Osorio Y, Ghiasi H. Comparison of adjuvant efficacy of herpes simplex virus type 1 recombinant viruses expressing TH1 and TH2 cytokine genes. J Virol 2003, 77: 5774–5783.
136 - Lee S, Gierynska M, Eo SK, Kuklin N, Rouse BT. Influence of DNA encoding cytokines on systemic and mucosal immunity following genetic vaccination against herpes simplex virus. Microbes Infect 2003, 5: 571–578.
137 - Veselenak RL, Shlapobersky M, Pyles RB, Wei Q, Sullivan SM, Bourne N. A Vaxfectin-adjuvanted HSV-2 plasmid DNA vaccine is effective for prophylactic and therapeutic use in the guinea pig model of genital herpes. Vaccine 2012, 30: 7046–7051. doi: 10.1016/j.vaccine.2012.09.057.
138 - Shlapobersky M, Marshak JO, Dong L, Huang ML, Wei Q, Chu A, Rolland A, Sullivan S, Koelle DM. Vaxfectin-adjuvanted plasmid DNA vaccine improves protection and immunogenicity in a murine model of genital herpes infection. J Gen Virol 2012, 93: 1305–1315. doi: 10.1099/vir.0.040055-0.
139 - Yan Y, Hu K, Deng X, Guan X, Luo S, Tong L, Du T, Fu M, Zhang M, Liu Y, Hu Q. Immunization with HSV-2 gB-CCL19 Fusion Constructs Protects Mice against Lethal Vaginal Challenge. J Immunol 2015, 195: 329–338. doi: 10.4049/jimmunol.1500198.
140 - Bright H, Perez DL, Christy C, Cockle P, Eyles JE, Hammond D, Khodai T, Lang S, West K, Loudon PT. The efficacy of HSV-2 vaccines based on gD and gB is enhanced by the addition of ICP27. Vaccine 2012, 30: 7529–7535. doi: 10.1016/j.vaccine.2012.10.046.
141 - Forrester A, Farrell H, Wilkinson G, Kaye J, Davis-Poynter N, Minson T. Construction and properties of a mutant of herpes simplex virus type 1 with glycoprotein H coding sequences deleted. J Virol 1992, 66: 341–348.
142 - McLean CS, Erturk M, Jennings R, Challanain DN, Minson AC, Duncan I, Boursnell ME, Inglis SC. Protective vaccination against primary and recurrent disease caused by herpes simplex virus (HSV) type 2 using a genetically disabled HSV-1. J Infect Dis 1994, 170: 1100–1109.
143 - Whitley RJ, Kern ER, Chatterjee S, Chou J, Roizman B. Replication, establishment of latency, and induced reactivation of herpes simplex virus gamma 1 34.5 deletion mutants in rodent models. J Clin Invest 1993, 91: 2837–2843.
144 - Brittle EE, Wang F, Lubinski JM, Bunte RM, Friedman HM. A replication-competent, neuronal spread-defective, live attenuated herpes simplex virus type 1 vaccine. J Virol 2008, 82: 8431–8441. doi: 10.1128/JVI.00551-08.
145 - Awasthi S, Zumbrun EE, Si H, Wang F, Shaw CE, Cai M, Lubinski JM, Barrett SM, Balliet JW, Flynn JA, Casimiro DR, Bryan JT, Friedman HM. Live attenuated herpes simplex virus 2 glycoprotein E deletion mutant as a vaccine candidate defective in neuronal spread. J Virol 2012, 86: 4586–4598. doi: 10.1128/JVI.07203-11.
146 - Keadle TL, Laycock KA, Morris JL, Leib DA, Morrison LA, Pepose JS, Stuart PM. Therapeutic vaccination with vhs(-) herpes simplex virus reduces the severity of recurrent herpetic stromal keratitis in mice. J Gen Virol 2002, 83: 2361–2365.
147 - Rajčáni J, Kúdelová M, Oravcová I, Vojvodová A, Košovský J, Matis J. Characterization of strain HSZP of herpes simplex virus type 1 (HSV1). Folia Microbiol (Praha) 1999, 44: 713–719.
148 - Meignier B. Genetically engineered attenuated herpes simplex viruses. Rev Infect Dis 1991, 13: S895–897.
149 - Halford WP, Püschel R, Gershburg E, Wilber A, Gershburg S, Rakowski B. A live-attenuated HSV-2 ICP0 virus elicits 10 to 100 times greater protection against genital herpes than a glycoprotein D subunit vaccine. PLoS One 2011, 6: e17748. doi: 10.1371/journal.pone.0017748.
150 - de Bruyn G, Vargas-Cortez M, Warren T, Tyring SK, Fife KH, Lalezari J, Brady RC, Shahmanesh M, Kinghorn G, Beutner KR, Patel R, Drehobl MA, Horner P, Kurtz TO, McDermott S, Wald A, Corey L. A randomized controlled trial of a replication defective (gH deletion) herpes simplex virus vaccine for the treatment of recurrent genital herpes among immunocompetent subjects. Vaccine 2006, 24: 914–920.
151 - Morrison LA, Knipe DM. Immunization with replication-defective mutants of herpes simplex virus type 1: sites of immune intervention in pathogenesis of challenge virus infection. J Virol 1994, 68: 689–696.
152 - Wachsman M, Kulka M, Smith CC, Aurelian L. A growth and latency compromised herpes simplex virus type 2 mutant (ICP10DeltaPK) has prophylactic and therapeutic protective activity in guinea pigs. Vaccine 2001, 19: 1879–1890.
153 - Da Costa XJ, Bourne N, Stanberry LR, Knipe DM. Construction and characterization of a replication-defective herpes simplex virus 2 ICP8 mutant strain and its use in immunization studies in a guinea pig model of genital disease. Virology 1997, 232: 1–12.
154 - Da Costa X, Kramer MF, Zhu J, Brockman MA, Knipe DM. Construction, phenotypic analysis, and immunogenicity of a UL5/UL29 double deletion mutant of herpes simplex virus 2. J Virol 2000, 74: 7963–7971.
155 - Delagrave S, Hernandez H, Zhou C, Hamberger JF, Mundle ST, Catalan J, Baloglu S, Anderson SF, DiNapoli JM, Londoño-Hayes P, Parrington M, Almond J, Kleanthous H. Immunogenicity and efficacy of intramuscular replication-defective and subunit vaccines against herpes simplex virus type 2 in the mouse genital model. PLoS One 2012, 7: e46714. doi: 10.1371/journal.pone.0046714.
156 - Brans R, Yao F. Immunization with a dominant-negative recombinant Herpes Simplex Virus (HSV) type 1 protects against HSV-2 genital disease in guinea pigs. BMC Microbiol 2010, 10: 163. doi: 10.1186/1471-2180-10-163.
157 - Casanova G, Cancela R, Alonzo L, Benuto R, Magana Mdel C, Hurley DR, Fishbein E, Lara C, Gonzalez T, Ponce R, Burnett JW, Calton GJ. A double-blind study of the efficacy and safety of the ICP10deltaPK vaccine against recurrent genital HSV-2 infections. Cutis 2002, 70: 235–239.
158 - van Lint AL, Torres-Lopez E, Knipe DM. Immunization with a replication-defective herpes simplex virus 2 mutant reduces herpes simplex virus 1 infection and prevents ocular disease. Virology 2007, 368: 227–231.
159 - Hoshino Y, Pesnicak L, Dowdell KC, Burbelo PD, Knipe DM, Straus SE, Cohen JI. Protection from herpes simplex virus (HSV)-2 infection with replication-defective HSV-2 or glycoprotein D2 vaccines in HSV-1-seropositive and HSV-1-seronegative guinea pigs. J Infect Dis 2009, 200: 1088–1095. doi: 10.1086/605645.
160 - Stanfield BA, Stahl J, Chouljenko VN, Subramanian R, Charles AS, Saied AA, Walker JD, Kousoulas KG. A single intramuscular vaccination of mice with the HSV-1 VC2 virus with mutations in the glycoprotein K and the membrane protein UL20 confers full protection against lethal intravaginal challenge with virulent HSV-1 and HSV-2 strains. PLoS One 2014, 9:e109890. doi: 10.1371/journal.pone.0109890.
161 - Zhang X, Castelli FA, Zhu X, Wu M, Maillère B, BenMohamed L. Gender-dependent HLA-DR-restricted epitopes identified from herpes simplex virus type 1 glycoprotein D. Clin Vaccine Immunol 2008, 15: 1436–1449. doi: 10.1128/CVI.00123-08.
162 - Mertz GJ, Peterman G, Ashley R, Jourden JL, Salter D, Morrison L, McLean A, Corey L. Herpes simplex virus type-2 glycoprotein-subunit vaccine: tolerance and humoral and cellular responses in humans. J Infect Dis 1984, 150: 242–249.
163 - Mertz GJ, Ashley R, Burke RL, Benedetti J, Critchlow C, Jones CC, Corey L. Double-blind, placebo-controlled trial of a herpes simplex virus type 2 glycoprotein vaccine in persons at high risk for genital herpes infection. J Infect Dis 1990, 161: 653–660
164 - Langenberg AG, Burke RL, Adair SF, Sekulovich R, Tigges M, Dekker CL, Corey L. A recombinant glycoprotein vaccine for herpes simplex virus type 2: safety and immunogenicity. Ann Intern Med 1995, 122: 889–898.
165 - Bernstein DI, Aoki FY, Tyring SK, Stanberry LR, St-Pierre C, Shafran SD, Leroux-Roels G, Van Herck K, Bollaerts A, Dubin G; GlaxoSmithKline Herpes Vaccine Study Group. Safety and immunogenicity of glycoprotein D-adjuvant genital herpes vaccine. Clin Infect Dis 2005, 40: 1271–1281.
166 - Stanberry LR, Spruance SL, Cunningham AL, Bernstein DI, Mindel A, Sacks S, Tyring S, Aoki FY, Slaoui M, Denis M, Vandepapeliere P, Dubin G; GlaxoSmithKline Herpes Vaccine Efficacy Study Group. Glycoprotein-D-adjuvant vaccine to prevent genital herpes. N Engl J Med 2002, 347: 1652–1661.
167 - Belshe RB, Leone PA, Bernstein DI, Wald A, Levin MJ, Stapleton JT, Gorfinkel I, Morrow RL, Ewell MG, Stokes-Riner A, Dubin G, Heineman TC, Schulte JM, Deal CD; Herpevac Trial for Women. Efficacy results of a trial of a herpes simplex vaccine. N Engl J Med 2012, 366: 34–43. doi: 10.1056/NEJMoa1103151.
168 - Tavares F, Cheuvart B, Heineman T, Arellano F, Dubin G. Meta-analysis of pregnancy outcomes in pooled randomized trials on a prophylactic adjuvanted glycoprotein D subunit herpes simplex virus vaccine. Vaccine 2013, 31: 1759–1764. doi: 10.1016/j.vaccine.2013.01.002.
169 - Cattamanchi A, Posavad CM, Wald A, Baine Y, Moses J, Higgins TJ, Ginsberg R, Ciccarelli R, Corey L, Koelle DM. Phase I study of a herpes simplex virus type 2 (HSV-2) DNA vaccine administered to healthy, HSV-2-seronegative adults by a needle-free injection system. Clin Vaccine Immunol 2008, 15: 1638–1643. doi: 10.1128/CVI.00167-08.
170 - Bernard MC, Barban V, Pradezynski F, de Montfort A, Ryall R, Caillet C, Londono-Hayes P. Immunogenicity, protective efficacy, and non-replicative status of the HSV-2 vaccine candidate HSV529 in mice and guinea pigs. PLoS One 2015, 10: e0121518. doi: 10.1371/journal.pone.0121518.
171 - Kutinová L, Benda R, Kalos Z, Dbalý V, Votruba T, Kvícalová E, Petrovská P, Doutlík S, Kamínková J, Domorázková E. Placebo-controlled study with subunit herpes simplex virus vaccine in subjects suffering from frequent herpetic recurrences. Vaccine 1988, 6: 223–228.
172 - Skinner GR, Turyk ME, Benson CA, Wilbanks GD, Heseltine P, Galpin J, Kaufman R, Goldberg L, Hartley CE, Buchan A. The efficacy and safety of Skinner herpes simplex vaccine towards modulation of herpes genitalis; report of a prospective double-blind placebo-controlled trial. Med Microbiol Immunol 1997, 186: 31–36.
173 - Straus SE, Corey L, Burke RL, Savarese B, Barnum G, Krause PR, Kost RG, Meier JL, Sekulovich R, Adair SF. Placebo-controlled trial of vaccination with recombinant glycoprotein D of herpes simplex virus type 2 for immunotherapy of genital herpes. Lancet 1994, 343: 1460–1463.
174 - Straus SE, Wald A, Kost RG, McKenzie R, Langenberg AG, Hohman P, Lekstrom J, Cox E, Nakamura M, Sekulovich R, Izu A, Dekker C, Corey L. Immunotherapy of recurrent genital herpes with recombinant herpes simplex virus type 2 glycoproteins D and B: results of a placebo-controlled vaccine trial. J Infect Dis 1997, 176: 1129–1134.
175 - Laing KJ, Dong L, Sidney J, Sette A, Koelle DM. Immunology in the Clinic Review Series; focus on host responses: T cell responses to herpes simplex viruses. Clin Exp Immunol 2012, 167: 47–58. doi: 10.1111/j.1365-2249.2011.04502.x
176 - Laing KJ, Magaret AS, Mueller DE, Zhao L, Johnston C, De Rosa SC, Koelle DM, Wald A, Corey L. Diversity in CD8(+) T cell function and epitope breadth among persons with genital herpes. J Clin Immun 2010, 30: 703–722. doi: 10.1007/s10875-010-9441-2.
177 - Berger C, Flowers ME, Warren EH, Riddell SR. Analysis of transgene-specific immune responses that limit the in vivo persistence of adoptively transferred HSV-TK-modified donor T cells after allogeneic hematopoietic cell transplantation. Blood 2006, 107: 2294–2302.
178 - Koelle DM, Liu Z, McClurkan CL, Cevallos RC, Vieira J, Hosken NA, Meseda CA, Snow DC, Wald A, Corey L. Immunodominance among herpes simplex virus-specific CD8 T cells expressing a tissue-specific homing receptor. Proc Natl Acad Sci U S A 2003, 100: 12899–12904.
179 - Chentoufi AA, Binder NR, Berka N, Durand G, Nguyen A, Bettahi I, Maillère B, BenMohamed L. Asymptomatic human CD4+ cytotoxic T-cell epitopes identified from herpes simplex virus glycoprotein B. J Virol 2008, 82: 11792–11802. doi: 10.1128/JVI.00692-08
180 - Wang X, Xie G, Liao J, Yin D, Guan W, Pan M, Li J, Li Y. Design and evaluation of a multi-epitope assembly peptide (MEAP) against herpes simplex virus type 2 infection in BALB/c mice. Virol J 2011, 8: 232. doi: 10.1186/1743-422X-8-232.
181 - Liu K, Jiang D, Zhang L, Yao Z, Chen Z, Yu S, Wang X. Identification of B- and T-cell epitopes from glycoprotein B of herpes simplex virus 2 and evaluation of their immunogenicity and protection efficacy. Vaccine 2012, 30: 3034–3041. doi: 10.1016/j.vaccine.2011.10.010.
182 - Muller WJ, Dong L, Vilalta A, Byrd B, Wilhelm KM, McClurkan CL, Margalith M, Liu C, Kaslow D, Sidney J, Sette A, Koelle DM. Herpes simplex virus type 2 tegument proteins contain subdominant T-cell epitopes detectable in BALB/c mice after DNA immunization and infection. J Gen Virol 2009, 90: 1153–1163. doi: 10.1099/vir.0.008771-0.
183 - Koelle DM, Frank JM, Johnson ML, Kwok WW. Recognition of herpes simplex virus type 2 tegument proteins by CD4 T cells infiltrating human genital herpes lesions. J Virol 1998, 72: 7476–7483.
184 - Novak EJ, Liu AW, Gebe JA, Falk BA, Nepom GT, Koelle DM, Kwok WW. Tetramer-guided epitope mapping: rapid identification and characterization of immunodominant CD4+ T cell epitopes from complex antigens. J Immunol 2001, 166: 6665–6670.
185 - Osorio Y, Mott KR, Jabbar AM, Moreno A, Foster TP, Kousoulas KG, Ghiasi H. Epitope mapping of HSV-1 glycoprotein K (gK) reveals a T cell epitope located within the signal domain of gK. Virus Res 2007, 128: 71–80.
186 - Mott KR, Chentoufi AA, Carpenter D, BenMohamed L, Wechsler SL, Ghiasi H. The role of a glycoprotein K (gK) CD8+ T-cell epitope of herpes simplex virus on virus replication and pathogenicity. Invest Ophthalmol Vis Sci 2009, 50: 2903–2912. doi: 10.1167/iovs.08-2957.
187 - Haynes JR, Arrington J, Dong L, Braun RP, Payne LG. Potent protective cellular immune responses generated by a DNA vaccine encoding HSV-2 ICP27 and the E. coli heat labile enterotoxin. Vaccine 2006, 24: 5016–5026.
188 - BenMohamed L, Bertrand G, McNamara CD, Gras-Masse H, Hammer J, Wechsler SL, Nesburn AB. Identification of novel immunodominant CD4+ Th1-type T-cell peptide epitopes from herpes simplex virus glycoprotein D that confer protective immunity. J Virol 2003, 77: 9463–9473.
189 - Cooper D, Mester JC, Guo M, Nasar F, Souza V, Dispoto S, Sidhu M, Hagen M, Eldridge JH, Natuk RJ, Pride MW. Epitope mapping of full-length glycoprotein D from HSV-2 reveals a novel CD4+ CTL epitope located at the transmembrane-cytoplasmic junction. Cell Immunol 2006, 239: 113–120.
190 - Chentoufi AA, Zhang X, Lamberth K, Dasgupta G, Bettahi I, Nguyen A, Wu M, Zhu X, Mohebbi A, Buus S, Wechsler SL, Nesburn AB, BenMohamed L. HLA-A*02:01-restricted CD8+ cytotoxic T lymphocyte epitopes identified from herpes simplex virus glycoprotein D. J Immunol 2008, 180: 426–437.
191 - Chentoufi AA, Dasgupta G, Christensen ND, Hu J, Choudhury ZS, Azeem A, Jester JV, Nesburn AB, Wechsler SL, BenMohamed L. A novel HLA (HLA-A*02:01) transgenic rabbit model for preclinical evaluation of human CD8+ T cell epitope-based vaccines against ocular herpes. J Immunol 2010, 184: 2561–2571. doi: 10.4049/jimmunol.0902322.
192 - Posavad CM, Remington M, Mueller DE, Zhao L, Magaret AS, Wald A, Corey L. Detailed characterization of T cell responses to herpes simplex virus-2 in immune seronegative persons. J Immunol 2010, 184: 3250–3259. doi: 10.4049/jimmunol.0900722.
193 - Posavad CM, Magaret AS, Zhao L, Mueller DE, Wald A, Corey L. Development of an interferon-gamma ELISPOT assay to detect human T cell responses to HSV-2. Vaccine 2011, 29: 7058–7066. doi: 10.1016/j.vaccine.2011.07.028.
194 - Maiers M, Gragert L, Klitz W. High-resolution HLA alleles and haplotypes in the United States population. Hum Immunol 2007, 68: 779–788.
195 - Koelle DM, Corey L. Herpes simplex: insights on pathogenesis and possible vaccines. Annu Rev Med 2008, 59: 381–395. doi: 10.1146/
196 - Samandary S, Kridane-Miledi H, Sandoval JS, Choudhury Z, Langa-Vives F, Spencer D, Chentoufi AA, Lemonnier FA, BenMohamed L. Associations of HLA-A, HLA-B and HLA-C alleles frequency with prevalence of herpes simplex virus infections and diseases across global populations: implication for the development of an universal CD8+ T-cell epitope-based vaccine. Hum Immunol 2014, 75: 715–729. doi: 10.1016/j.humimm.2014.04.016.
197 - Awasthi S, Friedman HM. Status of prophylactic and therapeutic genital herpes vaccines. Curr Opin Virol 2014, 6: 6–12. doi: 10.1016/j.coviro.2014.02.006.
198 - Wald A, Zeh J, Selke S, Warren T, Ryncarz AJ, Ashley R, Krieger JN, Corey L. Reactivation of genital herpes simplex virus type 2 infection in asymptomatic seropositive persons. N Engl J Med 2000, 342: 844–850.
199 - Khan AA, Srivastava R, Spencer D, Garg S, Fremgen D, Vahed H, Lopes PP, Pham TT, Hewett C, Kuang J, Ong N, Huang L, Scarfone VM, Nesburn AB, Wechsler SL, BenMohamed L. Phenotypic and functional characterization of herpes simplex virus glycoprotein B epitope-specific effector and memory CD8+ T cells from symptomatic and asymptomatic individuals with ocular herpes. J Virol 2015, 89: 3776–3792. doi: 10.1128/JVI.03419-14.