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Recent advances in mucosal vaccines and adjuvants Kristina Eriksson* and Jan Holmgren† Mucosal vaccines may be used both to prevent mucosal infections through the activation of antimicrobial immunity and to treat systemic inflammatory diseases through the induction of antigen-specific mucosal tolerance. New, efficient mucosal adjuvants for human use have been designed based on, amongst others, bacterial toxins and their derivatives, CpG-containing DNA, and different cytokines and chemokines, with the aim of improving the induction of mucosal Th1 and Th2 responses. Mucosal delivery systems, in particular virus-like particles, have been shown to enhance the binding, uptake and half-life of the antigens, as well as target the vaccine to mucosal surfaces. DNA vaccines are currently being developed for administration at mucosal surfaces. However, there have also been failures, such as the withdrawal of an oral vaccine against rotavirus diarrhea and a nasal vaccine against influenza, because of their potential side effects. Addresses Department of Medical Microbiology and Immunology and Göteborg University Vaccine Research Institute (GUVAX), Göteborg University, Guldhedsgatan 10A, 413 46 Göteborg, Sweden *e-mail: [email protected] † e-mail: [email protected] Current Opinion in Immunology 2002, 14:666–672 0952-7915/02/$ — see front matter © 2002 Elsevier Science Ltd. All rights reserved. DOI 10.1016/S0952-7915(02)00384-9 Abbreviations APC antigen presenting cell CpG ODN synthetic oligodeoxynucleotides containing immunostimulatory CpG motifs CT cholera toxin CTB cholera toxin B subunit CTL cytotoxic T lymphocyte DC dendritic cell DTH delayed type hypersensitivity FIV feline immunodeficiency virus GM-CSF granulocyte-macrophage colony-stimulating factor HSV herpes simplex virus Ig immunoglobulin IL interleukin ISCOM immune stimulating complex LT E. coli heat-labile enterotoxin LTB B subunit of LT M cell mucosal epithelial cell mLT detoxified LT adjuvant RSV respiratory syncytial virus SEA staphylococcal enteroxin A SIV simian immunodeficiency virus TGF transforming growth factor VLP virus-like particle

Introduction Mucosal immunization, especially by the oral route, has recently attracted much interest, both as a means of eliciting protective immunity against infectious diseases and as a possible approach for immunological treatment of diseases

that are caused by aberrant immune responses associated with tissue-damaging inflammation. The vast majority of infections occur at, or emanate from, mucosal surfaces. In these cases topical application of a vaccine is usually required to induce a protective immune response. Infections of this type include: infections of the gastrointestinal tract caused by Helicobacter pylori, Vibrio cholerae, enterotoxigenic Escherichia coli, Salmonella, Shigella spp., Campylobacter jejuni, Clostridium difficile, rotaviruses and calici viruses; infections of the respiratory tract caused by Mycoplasma pneumoniae, influenza virus and respiratory syncytial virus (RSV); infections of the urogenital tract such as those caused by HIV, Chlamydia, Neisseria gonorrhoeae and herpes simplex virus (HSV), and urinary tract infections caused by selected strains of E. coli. These infections still represent an enormous challenge for the development of vaccines that either prevent the infectious agent from attaching and colonizing the mucosal epithelium (noninvasive bacteria), or from penetrating and replicating within the mucosal epithelium (invasive bacteria and viruses), and/or can block the binding and action of microbial toxins. In most cases, although not in all, the main protective effector function elicited by mucosal immunization is the stimulation of secretory antimicrobial or antitoxic local immunoglobulin A (IgA) antibody responses and the associated mucosal immunologic memory. Mucosal immunization may also induce peripheralsystemic tolerance, especially against T cell-mediated immune reactions associated with the development of delayed type hypersensitivity (DTH) inflammatory reactions. This phenomenon, which is referred to as oral tolerance because it was initially documented as an effect of oral administration of antigen, is characterized by the fact that animals that have been fed with or have inhaled an antigen may become refractory or have diminished capacity to develop an immune response when re-exposed to the same antigen via parenteral injection. Oral tolerance is an important natural physiological mechanism whereby we avoid developing DTH and other allergic reactions to many ingested food proteins and other antigens. Indeed, certain cytokines, such as transforming growth factor (TGF)-β and IL-10, are known to be involved both in the production of secretory IgA at the mucosal surface and the induction of oral tolerance. As oral tolerance, in comparison to many other forms of immunotherapy, is specific for the initially ingested or inhaled antigen, and does not influence the development of systemic immune responses against other antigens, the induction of oral tolerance has become an attractive strategy for the prevention and potential treatment of illnesses that are caused by immunopathological reactions against specific foreign antigens and autoantigens. Therefore, mucosal immunization

Recent advances in mucosal vaccines and adjuvants Eriksson and Holmgren

and oral tolerance induction represent promising approaches to protect an individual against mucosal infectious agents and against systemic inflammatory ‘immunopathologies’ that are related to chronic infections, autoimmune disorders, and allergies. However, despite these attractive features, in practice it has often proven to be rather difficult to stimulate strong mucosal IgA immune responses by oral-mucosal administration of antigens, and the results of mucosal vaccination efforts using soluble protein antigens have been disappointing. Indeed, relatively few of the current vaccines that are approved for human use are administered mucosally: the oral polio vaccine, oral killed whole-cell B subunit and live-attenuated cholera vaccines, an oral live-attenuated typhoid vaccine, and an oral adenovirus vaccine (the latter vaccine being restricted to military personnel). Two recent additional mucosal vaccines, an oral live-attenuated vaccine against rotavirus diarrhea and a nasal enterotoxinadjuvanted inactivated influenza vaccine, were withdrawn after a short time on the market because of potential serious adverse reactions (intussusception and facial paresis, respectively), thus illustrating the complexity of mucosal vaccine development [1••]. Although promising results were obtained from initial clinical trials that utilized the principle of oral tolerance for immunotherapy of autoimmune diseases, such as multiple sclerosis and rheumatoid arthritis, extended randomized placebo-controlled multicenter trials failed to show any significant therapeutic benefits of these agents over those achieved by the placebo [2,3]. As will be discussed in this brief review, current efforts to overcome obstacles to the development of effective mucosal vaccines and/or immunotherapies are mainly directed towards finding a more efficient means of delivering appropriate antigens to the mucosal immune system, and towards discovering effective, safe mucosal adjuvants or immunoregulatory agents that provide protective immunity against infectious agents or induce the suppression of peripheral immunopathological disorders, respectively.

Mucosal adjuvants Bacterial toxins and derivatives

Although cholera toxin (CT) and the closely related E. coli heat-labile enterotoxin (LT) act as powerful mucosal adjuvants when co-administered with soluble antigens, their use in humans is hampered by their high toxicity. Both CT and LT consist of a homopentamer of cell-binding B subunits associated with a single toxic active A subunit. The A subunit enzymatically ribosylates the GS protein of adenylate cyclase and leads to increased cAMP production in the affected cells. Recently, site-directed mutagenesis has permitted the generation of LT and CT mutants that have reduced toxicity, but which retain significant adjuvanticity when given to animals by the nasal-mucosal route or, even though they then perform less well, by the oral-mucosal route [4]. Another approach that is used to circumvent the harmful drawbacks of CT or LT adjuvants is to link the

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enzymatically active A subunit domain of the toxin to a cell-binding moiety other than the natural B subunit, such as the cell-binding domain of Staphylococcus aureus protein A (CTA1–DD). CTA1–DD, like most other toxin derivatives, functions when applied nasally but not when given orally. This problem has recently been addressed by the incorporation of CTA1–DD fused to a short peptide into immune stimulating complexes (ISCOMS). These complexes are symmetrical colloidal particles with an open cage-like structure in the size range of 30–100 nm, composed of the saponin-adjuvant Quil A, cholesterol, phospholipids and the antigenic protein of interest. Oral vaccination with the ISCOM–CTA1–DD complex induced systemic and mucosal responses with both Th1 and Th2 characteristics [5•]. However, numerous (eight) immunizations were required, implying that the procedure needs further improvements. The B subunits of CT (CTB) and LT (LTB) can be used both as carrier molecules and as mucosal adjuvants, the efficacy of which in the latter case depends on the route of mucosal administration. Even though both CTB and LTB are poor adjuvants in animals when coadministered with noncoupled antigens by the oral route, both CTB and LTB have significant adjuvanticity via the nasal route. Thus, mice vaccinated with the influenza virus HA vaccine PR8 H1N1 together with LTB had higher levels of antiviral IgA and IgG, both in serum and in nasal and lung secretions, compared to mice that were given the subunit vaccine alone. The vaccinated mice were also protected against an intranasal viral challenge [6]. Similar results were obtained with a detoxified mutant of LT, called LT H44A [7]. CpG-containing DNA

Bacterial DNA containing cytosine-phosphate-guanosine (CpG) motifs, in which the cytosine is unmethylated, as well as synthetic oligodeoxynucleotides containing immunostimulatory CpG motifs (CpG ODN), have well-characterized adjuvant properties when administered systemically. The adjuvanticity of CpG DNA, which results from the binding of the CpG-rich DNA to the Toll-like receptor 9, is associated with the induction of both pro-inflammatory and Th1-inducing cytokines and chemokines, and the induction of MHC and costimulatory molecules on APCs. The resulting immune responses in mice are Th1 dominated with high levels of cytotoxic T lymphocytes (CTLs), IFN-γ production, and IgG2a antibody production. Recent studies show that CpG ODN are also effective mucosal adjuvants. Oral delivery of CpG ODN together with purified protein antigens promotes a mucosal Th2 response with IgA antibody formation and a systemic Th1 response [8]. The concomitant appearance of mucosal IgA antibodies that can prevent the uptake of mucosally transmitted pathogens in conjunction with systemic complement-activating antibodies and cytotoxic T cells would be of great benefit in combating most infections that take their departure from a mucosal surface. The applicability of this approach for inducing protective

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immunity was recently documented in a murine model of genital herpes infection. Nasal vaccination with a purified envelope glycoprotein (gB) from HSV-1 induced strong vaginal IgA and systemic IgG2a responses together with the induction of both systemic and mucosal CTLs. Importantly, the immune response obtained was crossprotective against the related HSV-2. Thus, approximately 50% of the animals were protected against a lethal vaginal challenge with HSV-2 [9•]. Cytokines and chemokines

Most adjuvants exert at least part of their adjuvant activity through the induction of inflammatory or Th1-inducing cytokines and chemokines. In general, the more potent the adjuvant, the more inappropriate it is for human use. One approach to circumvent the problem of overtly toxic adjuvants is to mimic the signals they induce in vivo, by simply adding these signaling molecules either directly as proteins, or indirectly as coding DNA. As mentioned above, the most powerful mucosal adjuvant identified to date is CT, which promotes strong mucosal IgA responses, systemic IgG responses and CTL responses to coadministered antigens. Several different combinations of cytokines can replace CT as nasal adjuvant. The most important of these is IL-1, which in combination with Th1-inducing cytokines such as IL-12, IL-18 and GM-CSF, can bring about mucosal and systemic responses with the same potency as CT [10]. The combination of IL-1 and IL-12/IL-18/GM-CSF also gives rise to a combined Th1 (CTL and IFN-γ) and Th2 (mucosal IgA) profile against less potent antigens, such as synthetic peptides [10]. The addition of genes that code for specific chemokines (for example, CCR7 ligands, which are involved in directing DCs to the T cell areas of secondary lymphoid organs) to a HSV-2 DNA plasmid vaccine produces an immunopotentiating effect in mice following either nasal or intragastric vaccination [11]. However, the protective efficacy of the chemokine-coding constructs against infection remains to be tested. The use of RANTES, which is a chemoattractant for monocytes, T cells and NK cells, and is a potent inducer of Th1 responses and CTLs, as a mucosal adjuvant appears to be promising. Nasal co-administration of RANTES with a protein antigen was shown to enhance Th1 and Th2 responses at both local and remote mucosal tissues, as well as systemically [12]. Whether chemokines are more attractive adjuvant candidates than chemokine receptors, or whether proteins are more effective in this respect than their corresponding coding DNA, remains to be determined. Recent studies of intranasal immunization of mice with CT, LT and related proteins that can bind to the GM1 ganglioside receptor used by both CT and LT, have shown that these proteins can, via retrograde transport, target olfactory tissues and redirect coadministered vaccine antigens to the same site [13••]. The significance of this finding for human applications remains to be determined. On one hand, there have been several clinical trials in which CTB

has been administered to humans without any signs of neurological or other serious adverse reactions. On the other hand, a nasal influenza vaccine, which was administered together with 2 µg of (fully toxic) LT, was recently withdrawn after several instances of facial paresis were noted among the vaccinees (C Spyr, personal communication). Furthermore, even though addition of LT or LT derivatives to a mucosal RSV vaccine induced strong CTL responses and protection against a viral challenge, the LT adjuvant was associated with enhanced disease due to the development of lung eosinophilia [14•]. Another approach to mucosal adjuvantation has been to use nontoxic bacterial products that target specific mucosal cells/tissues. The P40 outer membrane protein A of Klebsiella pneumoniae has proven to be an efficient carrier molecule for nasal vaccination, targeting CD11c+ mucosal DCs [15]. P40 fused or coupled to fragments from RSV promoted both systemic and mucosal immune responses and induced partial protection against a RSV challenge.

Mucosal DNA vaccination One of the most promising developments in vaccine research has been the use of nucleic acids as vaccines. DNA vaccination is based on the concept that the immunogenic moiety is made by the immunized vaccinee using the injected DNA as the template. DNA vaccines are easy and cheap to make, which adds to their popularity. DNA vaccination at mucosal surfaces has been tested extensively in the past two years with some promising results. Studies in mice that were given DNA intranasally [16] have shown rapid and relatively even distribution of plasmid DNA throughout the body. However, the DNA disappeared quickly from lymphoid organs and there was a somewhat alarming accumulation of plasmid DNA in the brain, from where it appeared to be cleared more slowly. Complications arise with mucosal DNA vaccination in the delivery and uptake of the DNA. The adsorption of DNA onto positively charged polylactide-co-glycolide microparticles permitted the efficient intranasal delivery of HIV-coding DNA, which induced a potent Th1-dominated systemic antiviral immune response [17]. Whether or not there is also a mucosal response in this case remains unknown. Another approach to circumvent the poor bioavailability of mucosally administered DNA has been to target the DNA to specialized mucosal epithelial cells, M cells, which line the inductive tissues of the gastrointestinal and nasal-respiratory tracts. It was reported that nasally delivered complexes of DNA and the M cell-binding σ1 protein from reovirus ( σ1 is the outer capsid cell attachment protein responsible for the selective and rapid attachment of reovirus to mucosal M cells, which represents the first crucial step in viral entry into the host) gave rise to strong antigen-specific mucosal IgA and IgG responses and, even more impressively, to specific CTLs in the lung that were directed against the encoded protein

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antigen, which demonstrated that both Th1 and Th2 effector cells accumulated within the mucous tissues [18•]. Although DNA vaccination is often highly efficacious in rodents, it has rarely worked so well in larger animals, including humans. Even though gene gun delivery of DNA intravulvomucosally into cattle primed for both systemic and mucosal T cell and B cell responses, it did not protect the animals against bovine herpesvirus 1 challenge [19]. Similarly, intradermal and rectal vaccination of macaques with a simian immunodeficiency virus (SIV) DNA vaccine gave rise to both systemic CTL responses and rectal IgA responses, but only two out of the nine monkeys were protected against a rectal viral challenge [20]. After oral delivery of a HIV DNA vaccine to humans, the vaccine induced local inflammation and activation of T cells within the mucosa [21], but negligible antigenspecific T cell and B cell responses. Thus, we conclude that the mucosal DNA vaccine field is still in its infancy, and that many improvements are needed to ensure efficacy in humans.

Mucosal delivery systems Great efforts have been made in recent years to combine mucosal delivery with agents that have intrinsic adjuvant activity. These systems include: microparticles that are based on lactide-co-glycolides; different types of lipid-based structures, such as liposomes, cochleates and ISCOMS; various live-attenuated bacteria and viruses; and mucosabinding lectins, such as the cell-binding B subunits of CT (CTB) or LT (LTB). Other developments are discussed below and include the use of commensal bacteria and viruslike particles as vectors for vaccine candidate antigens.

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viral transmission for vaccine delivery. Successful vaccination has been achieved in animals and humans with VLPs that were derived from several mucosal viral pathogens, including papillomavirus, Norwalk virus and hepatitis E virus. Oral delivery of hepatitis E VLPs was shown to be a strong inducer of systemic and mucosal IgA responses to both hepatitis E virus [23] and foreign epitopes that were expressed as chimeric proteins on the VLP surface [24]. Moreover, this induction of immunity did not require the co-administration of any external adjuvant. The intranasal delivery of VLPs appears to be more efficient and requires lower doses of antigen than oral delivery, at least in the case of the Norwalk agent VLPs [25•]. Interestingly, VLPs do not appear to function as adjuvants for co-delivered protein antigens [25•] but have adjuvanticity for both enclosed and admixed DNA vaccines [26•]. Thus, oral delivery of a papillomavirus pseudovirus containing DNA coding for a for a well-characterized CTL epitope induced strong antigen-specific CTL responses, whereas oral delivery of this construct alone did not induce a CTL response. Furthermore, vaccination with human papillomavirus VLPs coding for the papillomavirus oncogene E7 protected mice against mucosal challenge with an E7-expressing bovine papillomavirus VLP [26•], which supports the feasibility of this approach for mucosal vaccination. Another interesting VLP delivery system is the Sindbis virus replicon vector, which specifically targets immature DCs [27]. Both vaginal and rectal delivery of Sindbis virus replicon vectors expressing HIV gag proteins induced gag-specific IFN-γ-secreting T cells in the vaginal mucosa and protected the mice against vaginal challenge with a gag-expressing vaccinia virus [28•].

Commensal bacteria

Genital tract vaccination

Commensal bacteria represent an interesting delivery system for mucosal vaccines, in particular for vaccines that target diseases of the alimentary tract, as they can be manipulated to produce foreign antigens, they are adapted to the intestinal milieu, and they are both harmless and self-replicating. One example is Lactobacillus plantarum, which is normally found within the large bowel, but can be used for nasal vaccinations to induce both pulmonary and systemic immunity and to protect against a lethal challenge with tetanus toxin [22].

Nasal vaccination has emerged as the optimal vaccination route in rodents for the induction of genital antibody responses. The first study to investigate this approach in humans found that nasal vaccination was indeed a good way of inducing high titers of antibodies in vaginal (but not in cervical) secretions, whereas specific antibodies were detected at both these sites following vaginal vaccination [29]. However, the in vivo relevance of these differences in anatomical distribution of antibody remains to be determined. By contrast, nasal vaccination, as opposed to intramuscular vaccination, of rodents did not induce genital T cell responses or protection against genital Chlamydia trachomatis infection [30].

Virus-like particles

Pseudoviruses, or virus-like particles (VLPs), are selfassembling, nonreplicating viral core structures, often from non-enveloped viruses, which are produced recombinantly in vitro. As VLPs are cheap, easy to make and are highly immunogenic, there is considerable commercial interest in their use as viral vaccines. VLPs can also be used as antigenic carriers/adjuvants both for foreign antigens that are expressed recombinantly on their surface and for DNA vaccines that are carried within the VLPs. VLPs are especially interesting from a mucosal vaccine point of view, as they offer excellent opportunities to use the natural route of

Vaccine targeting to the genital draining lymph nodes was described several years ago as an efficient means of inducing local immunity against sexually transmitted diseases, for example, HIV/SIV. Anti- feline immunodeficiency virus (FIV) vaccines can now be added to this list, as targeted lymph node vaccination of cats with fixed FIV-infected cells protected against a later intrarectal cell-free inoculum of infectious FIV; the efficacy of this vaccine was poorer against cell-associated virus given via the same route [31].

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Despite its potential, we still believe that this deeply invasive route of vaccination, which requires ultrasound guidance, is inappropriate for large-scale human vaccination, particularly in poor and medically under-developed countries. However, an ingual-subcutaneous vaccination could possibly serve as a much easier and more acceptable means to achieve a similar response. A less invasive approach to STD vaccination that has shown promising results is intrarectal immunization using a synthetic SIV/HIV peptide vaccine together with a detoxified LT adjuvant (mLT). This mLT adjuvant, which was given rectally to macaques with the peptide vaccine, induced strong CTL responses both systemically and mucosally, particularly in combination with GM-CSF [32]. Repeated rectal vaccinations with peptides and mLT did not induce sterilizing immunity but protected the animals against persistent infection with a chimeric SIV/HIV virus [33]. The afforded protection obtained could be correlated with the activation of both CTLs and Th cells.

Immune deviation through mucosal antigen delivery Mucosal tolerance is a mechanism whereby the immune system refrains from responding in a deleterious manner to harmless substances that are contacted through mucosal surfaces, thereby permitting us to coexist with our normal flora and to eat large amounts of foreign food proteins without inducing harmful systemic immune responses. The immunological mechanisms behind this phenomenon are not fully understood, but at least three different mechanisms have been proposed: ignorance of the antigen by the immune system; the anergization or deletion of T cells that respond to the ingested antigen; and the generation of regulatory T cells that control and/or downmodulate the inflammatory response. Thus, various mucosally delivered ‘vaccines’ that mimic the natural induction of oral tolerance can be used as prophylactic or therapeutic treatments for inflammatory diseases. However, the application of this concept in patients with autoimmune diseases has largely failed to show any effect over that of placebo in large multi-center trials [2,3]. It appears clear that, just as adjuvants are required for effective induction of mucosal immunity, there is a need for delivery systems and/or adjuvants that enhance mucosal tolerance for effective immuntherapeutic application. Currently, the most promising vehicle for inducing mucosal tolerance is CTB. Recombinantly produced CTB, which was linked chemically or fused genetically to various autoantigens, has been shown to dramatically enhance the tolerogenic effect over that of mucosal administration of autoantigen alone, and to suppress the development of various autoimmune diseases in animal models [34]. It is now evident that mucosal administration of CTBconjugated antigens can also prevent infection-induced pathologic changes. Intranasal administration of conjugates of CTB and the egg antigen from Schistosoma mansoni

suppressed hepatic granuloma formation and reduced mortality following S. mansoni infection in mice (antipathologic immunity) in a process that involved the appearance of TGF-β-producing T cells. In addition, through an antibody-mediated effect, this treatment suppressed the worm burden and the production of parasite eggs (anti-infectious immunity) [35•]. CTB has also been shown to be antiinflammatory. Oral feeding of CTB prevented and cured Th1-driven experimental colitis in mice through an unidentified mechanism that involved reduced production of IL-12 within the large bowel [36•]. Mucosal tolerance/vaccination may also be used as a preventive treatment for superantigen-induced toxic shock. Nasal administration of small doses of staphylococcal enteroxin A (SEA) protected mice against a lethal systemic SEA challenge [37•]. The protection obtained was associated with significantly increased levels of IL-10 in the sera, indicating that regulatory T cells might be involved in the process.

Conclusion The development of mucosal vaccines, whether for prevention of infectious diseases or for immunotherapy of selected autoimmune, allergic or infectious-immunopathologic disorders, requires antigen delivery systems that can efficiently help to present vaccine or immunotherapy antigens to the mucosal immune system. Promising advances have been made in the design of both more efficient mucosal adjuvants and especially in the use of VLPs as mucosal vaccine delivery systems and CTB as carrier of antigens for immunotherapeutic tolerance induction. However, it is a memento that two recently developed mucosal vaccines for human use — against rotavirus diarrhea and influenza — were withdrawn after a short period because of adverse reactions among the vaccinees, emphasizing the difficult and challenging task of combining vaccine and adjuvant efficacy with safety and acceptability.

Acknowledgements We thank Vincent Collins for linguistic correction of the manuscript.

References and recommended reading Papers of particular interest, published within the annual period of review, have been highlighted as:

• of special interest •• of outstanding interest 1. ••

Murphy TV, Gargiullo PM, Massoudi MS, Nelson DB, Jumaan AO, Okoro CA, Zanardi LR, Setia S, Fair E, LeBaron CW et al.: Intussusception among infants given an oral rotavirus vaccine. N Engl J Med 2001, 344:564-572. A strong association between vaccination with the recently withdrawn tetravalent rhesus-human reassorted rotavirus vaccine and intussusception among otherwise healthy infants is shown. The authors predict that one case of intussusception would occur for each 5000–10000 infants vaccinated. 2.

Type 1 Diabetes Study Group: Effects of insulin in relatives of patients with type 1 diabetes mellitus. N Engl J Med 2002, 346:1685-1691.

3.

Choy EH, Scott DL, Kingsley GH, Thomas S, Murphy AG, Staines N, Panayi GS: Control of rheumatoid arthritis by oral tolerance. Arthritis Rheum 2001, 44:1993-1997.

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4.

Pizza M, Giuliani MM, Fontana MR, Monaci E, Douce G, Dougan G, Mills KH, Rappuoli R, Del Giudice G: Mucosal vaccines: non toxic derivatives of LT and CT as mucosal adjuvants. Vaccine 2001, 19:2534-2541.

5. •

17.

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Singh M, Vajdy M, Gardner J, Briones M, O’Hagan D: Mucosal immunization with HIV-1 gag DNA on cationic microparticles prolongs gene expression and enhances local and systemic immunity. Vaccine 2001, 20:594-602.

Mowat AM, Donachie AM, Jagewall S, Schon K, Lowenadler B, Dalsgaard K, Kaastrup P, Lycke N: CTA1-DD-immune stimulating complexes: a novel, rationally designed combined mucosal vaccine adjuvant effective with nanogram doses of antigen. J Immunol 2001, 167:3398-3405. A combined mucosal adjuvant delivery system, CTA1–DD–ISCOMS, consisting of a cell-binding moiety (the Ig-binding D moiety of S. aureus), an enzymatically active component (the A1 subunit of CT), and a mucosal delivery system (ISCOMS) is described, which is successfully used for both nasal and oral vaccination. In the latter case, however, the doses required are numerous.

18. Wu Y, Wang X, Csencsits KL, Haddad A, Walters N, Pascual DW: • M cell-targeted DNA vaccination. Proc Natl Acad Sci USA 2001, 98:9318-9323. A reovirus vaccine was targeted to the apical surface of M cells through the conjugation of the vaccine component(s) to polylysine. Intranasal vaccination of mice with this vaccine induced both mucosal IgA and CTL responses.

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Haan L, Verweij WR, Holtrop M, Brands R, van Scharrenburg GJ, Palache AM, Agsteribbe E, Wilschut J: Nasal or intramuscular immunization of mice with influenza subunit antigen and the B subunit of Escherichia coli heat-labile toxin induces IgA- or IgG-mediated protective mucosal immunity. Vaccine 2001, 19:2898-2907.

20. Wang SW, Kozlowski PA, Schmelz G, Manson K, Wyand MS, Glickman R, Montefiori D, Lifson JD, Johnson RP, Neutra MR et al.: Effective induction of simian immunodeficiency virus-specific systemic and mucosal immune responses in primates by vaccination with proviral DNA producing intact but noninfectious virions. J Virol 2000, 74:10514-10522.

7.

Hagiwar Y, Tsuji T, Iwasaki T, Kadowaki S, Asanuma H, Chen Z, Komase K, Suzuki Y, Aizawa C, Kurata T et al.: Effectiveness and safety of mutant Escherichia coli heat-labile enterotoxin (LT H44A) as an adjuvant for nasal influenza vaccine. Vaccine 2001, 19:2071-2079.

21. Lundholm P, Leandersson AC, Christensson B, Bratt G, Sandstrom E, Wahren B: DNA mucosal HIV vaccine in humans. Virus Res 2002, 82:141-145.

8.

McCluskie MJ, Weeratna RD, Krieg AM, Davis HL: CpG DNA is an effective oral adjuvant to protein antigens in mice. Vaccine 2000, 19:950-957.

9. •

Gallichan WS, Woolstencroft RN, Guarasci T, McCluskie MJ, Davis HL, Rosenthal KL: Intranasal immunization with CpG oligodeoxynucleotides as an adjuvant dramatically increases IgA and protection against herpes simplex virus- 2 in the genital tract. J Immunol 2001, 166:3451-3457. Intranasal vaccination of mice with HSV-1 glycoprotein B and CpG ODN adjuvant induced HSV-1/HSV-2 cross-protection and resistance to a genital HSV-2 challenge. The protection was associated with HSV-specific genital IgA, systemic CTL and mucosal CTL. 10. Staats HF, Bradney CP, Gwinn WM, Jackson SS, Sempowski GD, Liao HX, Letvin NL, Haynes BF: Cytokine requirements for induction of systemic and mucosal CTL after nasal immunization. J Immunol 2001, 167:5386-5394. 11. Eo SK, Lee S, Kumaraguru U, Rouse BT: Immunopotentiation of DNA vaccine against herpes simplex virus via co-delivery of plasmid DNA expressing CCR7 ligands. Vaccine 2001, 19:4685-4693. 12. Lilliard JWJ, Boyaka PN, Taub DD, McGhee JR: RANTES potentiates antigen-specific mucosal immune responses. J Immunol 2001, 166:162-169. 13. van Ginkel FW, Jackson RJ, Yuki Y, McGhee JR: Cutting edge: •• the mucosal adjuvant cholera toxin redirects vaccine proteins into olfactory tissues. J Immunol 2000, 165:4778-4782. The authors show that intranasally administered CT targets the olfactory bulb and redirects co-administered proteins to the same site. This study is particularly interesting in view of the recently withdrawn human nasal influenza vaccine. This particular vaccine was administered together with LT (which is closely related to CT), which was associated with several instances of facial paresis among the vaccinees.

19. Loehr BI, Willson P, Babiuk LA, van Drunen Littel-van den Hurk S: Gene gun-mediated DNA immunization primes development of mucosal immunity against bovine herpesvirus 1 in cattle. J Virol 2000, 74:6077-6086.

22. Grangette C, Muller-Alouf H, Goudercourt D, Geoffroy MC, Turneer M, Mercenier A: Mucosal immune responses and protection against tetanus toxin after intranasal immunization with recombinant Lactobacillus plantarum. Infect Immun 2001, 69:1547-1553. 23. Li T, Takeda N, Miyamura T: Oral administration of hepatitis E viruslike particles induces a systemic and mucosal immune response in mice. Vaccine 2001, 19:3476-3484. 24. Niikura M, Takamura S, Kim G, Kawai S, Saijo M, Morikawa S, Kurane I, Li TC, Takeda N, Yasutomi Y: Chimeric recombinant hepatitis E virus-like particles as an oral vaccine vehicle presenting foreign epitopes. Virology 2002, 293:273-280. 25. Guerrero RA, Ball JM, Krater SS, Pacheco SE, Clements JD, • Estes MK: Recombinant Norwalk virus-like particles administered intranasally to mice induce systemic and mucosal (fecal and vaginal) immune responses. J Virol 2001, 75:9713-9722. The authors show that VLPs are immunogenic when administered at a mucosal surface. Recombinant Norwalk VLPs given nasally to mice induced strong mucosal IgA responses in the absence of any external adjuvant. Oral vaccination was less promising. 26. Shi W, Liu J, Huang Y, Qiao L: Papillomavirus pseudovirus: a novel • vaccine to induce mucosal and systemic cytotoxic T-lymphocyte responses. J Virol 2001, 75:10139-10148. This paper shows that VLPs can be used as carriers/adjuvants for mucosal DNA vaccination. Oral vaccination with papillomavirus VLPs containing a DNA plasmid that encode for the regulatory papillomavirus protein E7 induced strong mucosal CTL responses and protected against challenge with an E7-expressing papillomavirus. 27.

Gardner JP, Frolov I, Perri S, Ji Y, MacKichan ML, zur Megede J, Chen M, Belli BA, Driver DA, Sherrill S et al.: Infection of human dendritic cells by a sindbis virus replicon vector is determined by a single amino acid substitution in the E2 glycoprotein. J Virol 2000, 74:11849-11857.

14. Simmons CP, Hussell T, Sparer T, Walzl G, Openshaw P, Dougan G: • Mucosal delivery of a respiratory syncytial virus CTL peptide with enterotoxin-based adjuvants elicits protective, immunopathogenic, and immunoregulatory antiviral CD8+ T cell responses. J Immunol 2001, 166:1106-1113. LT and a LT mutant lacking ADP-ribosyltransferase activity were shown to be strong adjuvants for intranasally administered RSV vaccines. The vaccination induced both RSV-specific CTL and protected against RSV challenge. However, administration of LT adjuvant was also associated with enhanced disease including lung eosinophilia.

28. Vajdy M, Gardner J, Neidleman J, Cuadra L, Greer C, Perri S, • O’Hagan D, Polo JM: Human immunodeficiency virus type 1 Gag-specific vaginal immunity and protection after local immunizations with sindbis virus-based replicon particles. J Infect Dis 2001, 184:1613-1616. Highly immunogenic, replication-defective, enveloped Sindbis virus vectors are described that can carry either foreign DNA and/or express foreign proteins. Vaginal or rectal vaccination of mice with sindbis-based replicon particles expressing HIV gag antigen protected mice against a vaginal challenge with vaccinia virus expressing the same HIV gag.

15. Goetsch L, Gonzalez A, Plotnicky-Gilquin H, Haeuw JF, Aubry JP, Beck A, Bonnefoy JY, Corvaia N: Targeting of nasal mucosaassociated antigen-presenting cells in vivo with an outer membrane protein A derived from Klebsiella pneumoniae. Infect Immun 2001, 69:6434-6444.

29. Johansson EL, Wassen L, Holmgren J, Jertborn M, Rudin A: Nasal and vaginal vaccinations have differential effects on antibody responses in vaginal and cervical secretions in humans. Infect Immun 2001, 69:7481-7486.

16. Oh YK, Kim JP, Hwang TS, Ko JJ, Kim JM, Yang JS, Kim CK: Nasal absorption and biodistribution of plasmid DNA: an alternative route of DNA vaccine delivery. Vaccine 2001, 19:4519-4525.

30. Igietseme JU, Murdin A: Induction of protective immunity against Chlamydia trachomatis genital infection by a vaccine based on major outer membrane protein-lipophilic immune responsestimulating complexes. Infect Immun 2000, 68:6798-6806.

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31. Finerty S, Stokes CR, Gruffydd-Jones TJ, Hillman TJ, Barr FJ, Harbour DA: Targeted lymph node immunization can protect cats from a mucosal challenge with feline immunodeficiency virus. Vaccine 2001, 20:49-58. 32. Belyakov IM, Ahlers JD, Clements JD, Strober W, Berzofsky JA: Interplay of cytokines and adjuvants in the regulation of mucosal and systemic HIV-specific CTL. J Immunol 2000, 165:6454-6462. 33. Belyakov IM, Hel Z, Kelsall B, Kuznetsov VA, Ahlers JD, Nacsa J, Watkins DI, Allen TM, Sette A, Altman J et al.: Mucosal AIDS vaccine reduces disease and viral load in gut reservoir and blood after mucosal infection of macaques. Nat Med 2001, 7:1320-1326. 34. Sun JB, Xiao BG, Lindblad M, Li BL, Link H, Czerkinsky C, Holmgren J: Oral administration of cholera toxin B subunit conjugated to myelin basic protein protects against experimental autoimmune encephalomyelitis by inducing transforming growth factor-beta-secreting cells and suppressing chemokine expression. Int Immunol 2000, 12:1449-1457. 35. Sun JB, Stadecker MJ, Mielcarek N, Lakew M, Li BL, Hernandez HJ, • Czerkinsky C, Holmgren J: Nasal administration of Schistosoma mansoni egg antigen-cholera B subunit conjugate suppresses

hepatic granuloma formation and reduces mortality in S. mansoni-infected mice. Scand J Immunol 2001, 54:440-447. The authors show that mucosal tolerance using CTB as a carrier protein can be used to treat infection-induced inflammatory pathology. Intranasal delivery of S. mansoni egg antigen conjugated to CTB reduced both liver granuloma formation and mortality in S. mansoni-infected mice. 36. Boirivant M, Fuss IJ, Ferroni L, De Pascale M, Strober W: Oral • administration of recombinant cholera toxin subunit B inhibits IL-12-mediated murine experimental (trinitrobenzene sulfonic acid) colitis. J Immunol 2001, 166:3522-3532. This study shows that CTB is a powerful inhibitor of Th1-driven immunopathology. Oral administration of CTB given either before or after onset of trinitrobenzene sulfonic acid (TNBS)-induced colitits prevented and resolved colitis, respectively. CTB treatment was associated with a marked inhibition of IL-12 secretion which prevented the development of a TNBSinduced Th1/IFN-γ response. 37. •

Collins LV, Eriksson K, Ulrich RG, Tarkowski A: Mucosal tolerance to a bacterial superantigen indicates a novel pathway to prevent toxic shock. Infect Immun 2002, 70:2282-2287. Mucosal vaccination with superantigen can also prevent superantigeninduced toxic shock syndrome. Intranasal administration of staphylococcal enterotoxin A (SEA) protected mice against a later lethal systemic SEA challenge. The protection was associated with enhanced levels of systemic IL-10.