Microbial products in allergy prevention and therapy.
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Animals↗
Biomedical subjects
E Raz
Publications and source records attributed to E Raz.
Immunostimulatory DNA-based therapeutics for experimental and clinical allergy.
Although allergen immunotherapy (IT) has been shown to modulate allergic hypersensitivities, its efficacy is limited. Recently, in various models of experimental allergy, a number of reagents which we have termed immunostimulatory DNA-based therapeutics have proven highly effective in both the prevention and reversal of Th2 mediated hypersensitivity states. These include immunization with gene vaccines, allergen mixed with immunostimulatory oligodeoxynucleotide (ISS-ODN), and physical allergen: ISS-conjugates (AIC), and immunomodulation with ISS-ODN alone. Results from our laboratory have shown that immunostimulatory DNA-based therapeutics may be effective for the reversal of allergic hypersensitivity states in humans and several clinical trials have already been initiated. This review will focus on our laboratory's experience with immunostimulatory DNA-based therapeutics in various murine models of allergy and their potential utility in the treatment of allergic patients.
Animals↗
A zebrafish nanos-related gene is essential for the development of primordial germ cells.
Asymmetrically distributed cytoplasmic determinants collectively termed germ plasm have been shown to play an essential role in the development of primordial germ cells (PGCs). Here, we report the identification of a nanos-like (nanos1) gene, which is expressed in the germ plasm and in the PGCs of the zebrafish. We find that several mechanisms act in concert to restrict the activity of Nanos1 to the germ cells including RNA localization and control over the stability and translatability of the RNA. Reducing the level of Nanos1 in zebrafish embryos revealed an essential role for the protein in ensuring proper migration and survival of PGCs in this vertebrate model organism.
Amino Acid Sequence↗
Identification of immunostimulatory DNA-induced genes by suppression subtractive hybridization.
Bacterial DNA and related synthetic immunostimulatory oligodeoxyribo-nucleotides (ISS-ODN) have stimulatory effects on mammalian immune cells through a Toll-like receptor, TLR9. Genes upregulated in ISS-ODN-stimulated immune cells are obviously significant to delineate the mechanism of the induced innate immunity. Employing suppression subtractive hybridization (SSH), we have generated a profile of genes induced by ISS-ODN in spleen cells. Sequencing of 87 clones isolated by the SSH showed 39 clones corresponding to known mouse genes in the public database. Eleven clones appeared to possess 80-90% homology with known mouse genes and the remaining 37 clones showed no significant homology with any known mouse genes. A series of known genes which have not previously been reported to be induced with ISS-ODN were confirmed to be induced in ISS-ODN-stimulated bone marrow-derived macrophages: NF-kappaB p105, IRF-1, PA28beta, IRG2, and MyD88. These genes were suggested to be involved in the molecular process of innate host defense mechanisms.
Adjuvants, Immunologic↗
Immunostimulatory DNA-based vaccines elicit multifaceted immune responses against HIV at systemic and mucosal sites.
Immunostimulatory DNA sequences (ISS, also known as CpG motifs) are pathogen-associated molecular patterns that are potent stimulators of innate immunity. We tested the ability of ISS to act as an immunostimulatory pathogen-associated molecular pattern in a model HIV vaccine using gp120 envelope protein as the Ag. Mice immunized with gp120 and ISS, or a gp120:ISS conjugate, developed gp120-specific immune responses which included: 1) Ab production; 2) a Th1-biased cytokine response; 3) the secretion of beta-chemokines, which are known to inhibit the use of the CCR5 coreceptor by HIV; 4) CTL activity; 5) mucosal immune responses; and 6) CD8 T cell responses that were independent of CD4 T cell help. Based on these results, ISS-based immunization holds promise for the development of an effective preventive and therapeutic HIV vaccine.
AIDS Vaccines↗
Induction of antigen-specific Th1-biased immune responses by plasmid DNA in schistosoma-infected mice with a preexistent dominant Th2 immune profile.
A requisite for vaccines to confer protection against intracellular infections such as Human Immunodeficiency Virus or Mycobacterium tuberculosis is their capacity to induce Th1 immune responses. However, they may fail to do so in Africa and South East Asia, where most individuals have a dominant preexistent Th2 immune profile, due to persistent helminthic parasitic infections, which may undermine any Th1 response. It is well established that DNA vaccines induce strong Th1 biased immune responses against an encoded antigen, depending on the route and mode of immunization. Here, we demonstrate that intradermal immunization with plasmid DNA encoding beta-gal (pCMV-LacZ) of Schistosoma-infected mice, with preexistent dominant Th2 immune background, induce a strong Th1 anti-beta-gal response, as opposed to immunized with beta-gal only. Importantly, the established protective Th2 immune response to schistosomes was not disrupted. These findings strongly support the possibility of using plasmid DNA as a Th1 inducing adjuvant when immunizing populations with a strong preexistent Th2 immune profile.
Animals↗
Type I interferon is required to mount an adaptive response to immunostimulatory DNA.
Immunostimulatory DNA sequences (ISS, CpG motifs) potently stimulate Th1 and cytotoxic T lymphocyte (CTL) responses to antigens and have thus generated considerable interest due to their potential use in immunotherapeutics. An array of cytokines are produced in response to ISS exposure, but the relative importance of each of these mediators in the stimulation of innate and adaptive ISS-induced immunity has yet to be fully investigated. To address this issue, we measured immune responses in mice with targeted deletions of the ISS-induced genes encoding IL-12 (IL-12(-/-)), IFN-gamma (IFN-gamma(-/-)), the IFN-gamma receptor (IFN-gammaR(-/-)), and the IFN-alpha/beta receptor (IFN-alpha/betaR(-/-)) after immunization with ISS-containing oligodeoxynucleotides and model antigens. IL-12(-/-) and IFN-alpha/betaR(-/-) mice were compromised in their ability to develop a cross-primed CTL response, whereas IFN-gamma(-/-) and IFN-gammaR(-/-) mice were not. In addition, lymphocytes from immunized IFN-alpha/betaR(-/-) mice had defective IFN-gamma responses to antigen restimulation. Antigen nonspecific ISS-induced B cell proliferation was normal in the four deficient strains; however, innate IL-6 production was reduced in IFN-gamma(-/-) and IFN-gammaR(-/-) splenocytes and eliminated in IFN-alpha/betaR(-/-) cells. While IL-12 production was defective in only the IFN-gamma(-/-) splenocytes, innate natural killer cell IFN-gamma synthesis was virtually absent in the IL-12(-/-) and IFN-alpha/betaR(-/-) mice. Thus, while IFN-alpha/beta, IFN-gamma, and IL-12 each play important and distinct roles in the development of the innate and adaptive immune responses to ISS, IFN-alpha/beta is a particularly crucial and currently under-appreciated factor in this system.
Adjuvants, Immunologic↗
Systemic administration of immunostimulatory DNA sequences mediates reversible inhibition of Th2 responses in a mouse model of asthma.
This study investigated whether immunostimulatory DNA sequences (ISS) induce a transient or sustained inhibition of Th2 responses to inhaled antigen. We sensitized mice with subcutaneous injections to develop a Th2 response to ovalbumin (ova) and then administered a dose of ISS prior to ova inhalation challenge. Mice were then rechallenged with ova by inhalation a second time at varying time points after the first ova inhalation (1 to 8 weeks later) to determine whether the ISS dose administered prior to the first ova inhalation protected against a subsequent second ova inhalation challenge. A single dose of ISS inhibited the Th2 response to the first inhalation of ova antigen, as well as 4 weeks later to the second inhalation of ova. However, ISS did not inhibit a Th2 response to the second inhalation of ova 8 weeks later. The reversible inhibition of Th2 responses at 8 weeks suggests the need for repeated ISS administration at monthly intervals.
Adjuvants, Immunologic↗
DNA-based immunotherapeutics for the treatment of allergic disease.
Allergic diseases are a growing health concern in industrialized countries. Despite a number of effective therapeutic options for the prevention and treatment of the pathophysiologic responses which characterize allergic diseases, the induction of true allergen desensitization remains an elusive therapeutic goal. Only immunotherapy (IT) has been shown to have any effect on the underlying hypersensitivities which mediate allergic reactions, and traditional protein-based allergen IT has a limited scope of efficacy However, a number of reagents collectively termed DNA-based immunotherapeutics have proven highly effective in both the prevention and reversal of Th2-mediated hypersensitivity states in mouse models of allergic disease. Four basic DNA-based immunotherapeutic modalities have been used for these studies. These include immunization with gene vaccines, allergen mixed with immunostimulatory oligodeoxynucleotide (ISS-ODN), and physical allergen-ISS-ODN conjugates (AIC), as well as immunomodulation with ISS-ODN alone. Results from many laboratories have generated guarded optimism that DNA-based immunotherapeutics may be effective for the reversal of allergic hypersensitivity states in humans, and several clinical trials have already been initiated. This review will focus on our present understanding of the biological activities of DNA-based immunotherapeutics and their application to the treatment of allergic diseases.
Adjuvants, Immunologic↗
Immunostimulatory DNA inhibits IL-4-dependent IgE synthesis by human B cells.
BACKGROUND: Immunostimulatory sequence oligodeoxynucleotide (ISS-ODN) is a potent antiallergic immunomodulating agent in mice. However, few studies have addressed its antiallergic potential in human subjects. OBJECTIVE: We sought to determine whether a phosphoro-thioate ISS-ODN could inhibit IL-4-dependent IgE synthesis by human B cells. METHODS: Initially, nonatopic- and atopic-donor PBMCs were incubated with ISS-ODN or mutated oligodeoxynucleotide, and cytokine production and B-cell expression of IFN-gamma receptor and IL-4 receptor were measured by using ELISA and flow cytometry, respectively. In subsequent studies atopic-donor PBMCs were incubated with IL-4 alone or with ISS-ODN or mutated oligodeoxynucleotide. After 14 days, IgE production and IgM, IgG, and IgA production were determined by using ELISA. In select IgE studies cytokines were neutralized with mAbs. RESULTS: ISS-ODN induced IL-12, IFN-alpha, IFN-gamma, IL-10, and IL-6 production from both nonatopic- and atopic-donor PBMCs. ISS-ODN also increased IFN-gamma receptor and inhibited IL-4 receptor expression on B cells from both donor populations. Furthermore, ISS-ODN inhibited IL-4-dependent IgE production by atopic-donor PBMCs. Neutralization of IL-12, IFN-alpha, IFN-gamma, and IL-10, but not IL-6, attenuated the inhibitory activity of ISS-ODN on IgE production. In contrast to its inhibition of IgE synthesis, ISS-ODN stimulated the production of IgM, IgG, and IgA. CONCLUSION: These in vitro studies demonstrate that phos-phorothioate ISS-ODN elicits an innate immune response by PBMCs, which inhibits IL-4-dependent IgE synthesis. In addition, these results provide further support for consideration of ISS-ODN therapy for the treatment of allergic disease in clinical practice.
Anti-Allergic Agents↗
Immunostimulatory DNA mediates inhibition of eosinophilic inflammation and airway hyperreactivity independent of natural killer cells in vivo.
BACKGROUND: Immunostimulatory DNA sequences (ISS) inhibit eosinophilic inflammation and airway hyperreactivity in mouse models of asthma. In vitro ISS activate natural killer (NK) cells to secrete IFN-gamma, and this cytokine is hypothesized to contribute to the antiallergic effect of ISS in vivo. OBJECTIVE: We investigated whether ISS activation of NK cells is important in mediating the reduction in airway hyperreactivity and the antieosinophilic effect of ISS in vivo. METHODS: We assessed whether ISS modulated the development of eosinophilic airway inflammation and airway hyperreactivity to methacholine in ovalbumin (OVA)-sensitized and OVA allergen-challenged mice pretreated with an antibody to deplete NK cells. RESULTS: Mice sensitized and challenged with OVA had significant bronchoalveolar lavage and lung eosinophilia, as well as airway hyperresponsiveness. ISS induced significant inhibition of bronchoalveolar lavage and lung eosinophilia, as well as airway hyperresponsiveness, in OVA-sensitized mice pretreated before OVA challenge with an NK cell-depleting antibody (NK(-) mice), as well as in mice pretreated with a control non-NK cell-depleting antibody (NK(+) mice). The NK cell-depleting antibody inhibited ISS-induced IFN-gamma production by spleen cells. CONCLUSION: These studies demonstrate that depletion of NK cells has no significant effect on ISS-mediated inhibition of airway eosinophilia and airway hyperresponsiveness in vivo, suggesting that non-NK cells and cytokines other than IFN-gamma derived from NK cells mediate the majority of the ISS-inhibitory effect on eosinophilic inflammation and airway hyperresponsiveness in vivo.
Adjuvants, Immunologic↗
Immunostimulatory DNA sequences inhibit respiratory syncytial viral load, airway inflammation, and mucus secretion.
BACKGROUND: Immunostimulatory DNA sequences (ISS) activate the innate immune system to generate antiviral cytokines, such as IFN-gamma. OBJECTIVE: This study investigated whether ISS could reduce viral load, mucus secretion, airway inflammation, and airway hyperreactivity to methacholine in a mouse model of respiratory syncytial virus (RSV) infection. METHODS: Mice were pretreated with ISS 6 days before RSV infection, and lung indices of RSV viral load (viral titer and PCR), bronchoalveolar lavage fluid cytokines (IFN-gamma), airway inflammation (peribronchial inflammation and periodic acid-Schiff-positive mucus cells), and airway hyperreactivity (methacholine responsiveness) were assessed 4 to 6 days after RSV infection. RESULTS: ISS induced the expression of the antiviral cytokine IFN-gamma in the lung, and this was associated with significantly reduced RSV viral titers, mucus secretion, and peribronchial inflammation. ISS reduced, but did not significantly inhibit, RSV-induced airway hyperreactivity to methacholine. CONCLUSION: Because ISS induced significant levels of lung IFN-gamma, an immunization strategy based solely on the administration of IFN-gamma may be insufficient to inhibit RSV-induced airway hyperreactivity to methacholine, an endpoint important in the subset of RSV-infected subjects with asthma.
Adjuvants, Immunologic↗
Enhancement of innate immunity against Mycobacterium avium infection by immunostimulatory DNA is mediated by indoleamine 2,3-dioxygenase.
Bacterial DNA and its synthetic immunostimulatory oligodeoxynucleotide analogs (ISS-ODN) activate innate immunity and promote Th1 and cytotoxic T-lymphocyte immune responses. Based on these activities, we investigated whether ISS-ODN could modify the course of Mycobacterium avium infection. M. avium growth in vitro was significantly inhibited by ISS-ODN treatment of human and mouse macrophages, and M. avium growth in vivo was similarly inhibited in C57BL/6 mice treated with ISS-ODN. This protective effect of ISS-ODN was largely independent of tumor necrosis factor alpha (TNF-alpha), interleukin 12 (IL-12), nitric oxide, NADPH oxidase, alpha/beta interferon (IFN-alpha/beta), and IFN-gamma. In contrast, we found that the induction of indoleamine 2,3-dioxygenase (IDO) was required for the antimycobacterial effect of ISS-ODN. To evaluate the potential for synergism between ISS-ODN and other antimycobacterial agents, treatment with a combination of ISS-ODN and clarithromycin (CLA) was tested in vitro and in vivo. ISS-ODN significantly enhanced the therapeutic effect of CLA in both human and mouse macrophages and in C57BL/6 mice. This study newly identifies IDO as being involved in the antimicrobial activity of ISS-ODN and suggests the usefulness of ISS-ODN when used in combination with conventional chemotherapy for microbial infections.
Adjuvants, Immunologic↗
DNA-PKcs is required for activation of innate immunity by immunostimulatory DNA.
Bacterial DNA and related synthetic immunostimulatory oligodeoxyribonucleotides (ISS-ODN) stimulate innate immunity. However, the molecular recognition mechanism that initiates signaling in response to bacterial DNA and ISS-ODN has not been identified. Herein, we demonstrate that administration of bacterial DNA and ISS-ODN to mice lacking the catalytic subunit of DNA-PK (DNA-PKcs) and in vitro stimulation of BMDM from these mice result in defective induction of IL-6 and IL-12. Further analysis using BMDM of IKKbeta(-/-) revealed that both DNA-PKcs and IKKbeta are essential for normal cytokine production in response to ISS-ODN or bacterial DNA. ISS-ODN and bacterial DNA activate DNA-PK, which in turn contributes to activation of IKK and NF-kappaB. These results reveal a novel role of DNA-PKcs in innate immune responses and a link between DNA repair and innate immunity.
Androstadienes↗
The function and regulation of vasa-like genes in germ-cell development.
The vasa gene, essential for germ-cell development, was originally identified in Drosophila, and has since been found in other invertebrates and vertebrates. Analysis of these vasa homologs has revealed a highly conserved role for Vasa protein among different organisms, as well as some important differences in its regulation.
Animals↗
Systemic or mucosal administration of immunostimulatory DNA inhibits early and late phases of murine allergic conjunctivitis.
Seasonal allergic conjunctivitis is one of the most common manifestations of allergic disease, affecting 15 % population in the United States annually. Short ragweed (RW) is a major cause of seasonal allergies. Immunostimulatory DNA sequences (ISS or CpG motifs) can inhibit an on-going Th2/allergic response and induce a de novo Th1 response. In this study, we investigated the ability of these ISS to modulate allergic responses in a RW-induced mouse model of seasonal allergic conjunctivitis. Systemic or mucosal administration of ISS oligonucleotide (ISS-ODN) after RW sensitization inhibited both the immediate hypersensitivity response and the late-phase cellular infiltration and induced a RW-specific Th1 response. ISS-ODN administration suppressed the rise of RW-specific IgE titers after repeated allergen challenge. Furthermore, ISS administration was more effective than dexamethasone in inhibiting the allergic response. Mechanistically, the ISS-induced immunomodulatory effects were abolished when mice were treated with anti-IL-12 neutralizing antibodies, suggesting a pivotal role for type 1 cytokines in the inhibition of both the immediate hypersensitivity and the late-phase cellular infiltration. Thus, ISS-ODN is a novel anti-inflammatory and immunomodulatory agent that significantly inhibits the allergic response and may provide an alternative to the current standard care of ocular allergy.
Administration, Topical↗
Conjugation of protein to immunostimulatory DNA results in a rapid, long-lasting and potent induction of cell-mediated and humoral immunity.
Immunostimulatory DNA sequences (ISS) are a potent Th1 adjuvant. We hypothesized that conjugation of ISS to protein antigens would strongly enhance their immunogenicity because both antigen and adjuvant (ISS) would be delivered to the same locale/antigen-presenting cell. To test this hypothesis, we conjugated a 22-mer immunostimulatory oligodeoxynucleotide (ISS-ODN) to two test antigens of differing intrinsic immunogenicity, namely Escherichia coli beta-galactosidase and the HIV-1 envelope glycoprotein gp120. We show that the antigen-ISS conjugates rapidly induce Th1 cells secreting high levels of IFN-gamma, strong CTL activity, and high titer IgG2a and HIV-neutralizing antibodies, exceeding gene and protein vaccination alone or immunization with mixtures of antigen and ISS-ODN. The data suggest that this procedure generates a novel and unique vaccine that rapidly triggers strong humoral and cell-mediated immunity.
Animals↗
Immunostimulatory sequence oligodeoxynucleotide: A novel mucosal adjuvant.
The majority of infectious agents enter the body via mucosal surfaces. Therefore, there is great interest in developing vaccination strategies that lead to mucosal immunity. However, to generate strong mucosal immune responses to nonreplicating immunogens, mucosal delivery with an adjuvant is required, and no safe and effective mucosal adjuvants are presently available for use in humans. Recently, immunostimulatory sequence oligodeoxynucleotide (ISS-ODN) has been shown to be a potent mucosal adjuvant. Intranasal (i.n.) delivery of antigen with ISS-ODN elicits a secretory IgA response both locally in respiratory secretions and distally in intestinal and vaginal secretions. In this regard, ISS-ODN has been shown to be as effective as cholera toxin (CT), but unlike CT, ISS-ODN has not been shown to have significant toxicity. Furthermore, i.n. antigen/ISS-ODN coimmunization induces a strong Th(1)-biased systemic immune response with cytotoxic T lymphocyte activity. These observations suggest that ISS-ODN would be an ideal mucosal adjuvant for development of vaccines against mucosal pathogens.
Adjuvants, Immunologic↗