Induction of cellular and molecular Immunomodulatory pathways by vitamin E and vitamin C.

INTRODUCTION: Vitamins E and C are well known small molecules that have been used to maintain health for decades. Recent studies of the cellular and molecular pathways leading to immunomodulation by these molecules have been of interest, as have their anti-oxidant properties and signal transduction pathways for curing or improving infectious diseases and cancer. Areas covered: Herein, the authors provide a definition and the structural classification of vitamins E and C and how these molecules influence cellular function. The studies include in vitro, ex vivo and in vivo studies in animal models as well as clinical trials. The authors give particular focus to the scientifically factual and putative roles of these molecules in innate and adaptive immunomodulation and prevention or cure of diseases. Expert opinion: The antioxidant properties of vitamins E and C are well studied. However, whether there is a link between their antioxidant and immunomodulation properties is unclear. In addition, there is a strong, albeit putative, prevailing notion that vitamin C can prevent or cure infectious diseases or cancer. Presently, while there is proven evidence that vitamin E possesses immunomodulatory properties that may play a positive role in disease outcomes, this evidence is less available for vitamin C.

Comparative Safety and Efficacy Profile of a Novel Oil in Water Vaccine Adjuvant Comprising Vitamins A and E and a Catechin in Protective Anti-Influenza Immunity.

Non-replicating vaccines, such as those based on recombinant proteins, require adjuvants and delivery systems, which have thus far depended on mimicking pathogen danger signals and strong pro-inflammatory responses. In search of a safer and more efficacious alternative, we tested whether vaccinations with influenza recombinant hemagglutinin (HA) mixed with a novel vegetable oil in water emulsion adjuvant (Natural Immune-enhancing Delivery System, NIDS), based on the immune-enhancing synergy of vitamins A and E and a catechin, could protect against intra-nasal challenge with live influenza virus. Vaccinations of inbred Brag Albino strain c (BALB/c) mice, with HA mixed with NIDS compared to other adjuvants, i.e., a squalene oil in water emulsion (Sq. oil), and the Toll Like Receptor 3 (TLR3) agonist Poly (I:C), induced significantly lower select innate pro-inflammatory responses in serum, but induced significantly higher adaptive antibody and splenic T Helper 1 (TH1) or TH2, but not TH17, responses. Vaccinations with NIDS protected against infection, as measured by clinical scores, lung viral loads, and serum hemagglutination inhibition titers. The NIDS exhibited a strong dose sparing effect and the adjuvant action of NIDS was intact in the outbred CD1 mice. Importantly, vaccinations with the Sq. oil, but not NIDS, induced a significantly higher Serum Amyloid P component, an acute phase reactant secreted by hepatocytes, and total serum IgE. Thus, the NIDS may be used as a clinically safer and more efficacious vaccine adjuvant against influenza, and potentially other infectious diseases.

Vitamin A or E and a catechin synergize as vaccine adjuvant to enhance immune responses in mice by induction of early interleukin-15 but not interleukin-1ß responses.

Vitamins A and E and select flavonoids in the family of catechins are well-defined small molecules that, if proven to possess immunomodulatory properties, hold promise as vaccine adjuvants and various therapies. In an effort to determine the in vivo immunomodulatory properties of these molecules, we found that although mucosal and systemic vaccinations with a recombinant HIV-1BaL gp120 with either a catechin, epigallo catechin gallate (EGCG) or pro-vitamin A (retinyl palmitate) alone in a vegetable-oil-in-water emulsion (OWE) suppressed antigen-specific responses, the combination of EGCG and vitamin A or E in OWE (Nutritive Immune-enhancing Delivery System, NIDS) synergistically enhanced adaptive B-cell, and CD4(+) and CD8(+) T-cell responses, following induction of relatively low local and systemic innate tumour necrosis factor-α (TNF-α), interleukin-6 (IL-6) and IL-17, but relatively high levels of early systemic IL-15 responses. For induction of adaptive interferon-γ and TNF-α responses by CD4(+) and CD8(+) T cells, the adjuvant effect of NIDS was dependent on both IL-15 and its receptor. In addition, the anti-oxidant activity of NIDS correlated positively with higher expression of the superoxide dismutase 1, an enzyme involved in reactive oxygen species elimination but negatively with secretion of IL-1ß. This suggests that the mechanism of action of NIDS is dependent on anti-oxidant activity and IL-15, but independent of IL-1ß and inflammasome formation. These data show that this approach in nutritive vaccine adjuvant design holds promise for the development of potentially safer effective vaccines.

Induction of cellular and molecular immunomodulatory pathways by vitamin A and flavonoids.

INTRODUCTION: A detailed study of reports on the immunomodulatory properties of vitamin A and select flavonoids may pave the way for using these natural compounds or compounds with similar structures in novel drug and vaccine designs against infectious and autoimmune diseases and cancers. AREAS COVERED: Intracellular transduction pathways, cellular differentiation and functional immunomodulatory responses have been reviewed. The reported studies encompass in vitro, in vivo preclinical and clinical studies that address the role of vitamin A and select flavonoids in induction of innate and adaptive B- and T-cell responses, including TH1, TH2 and regulatory T cells (Treg). EXPERT OPINION: While the immunomodulatory role of vitamin A, and related compounds, is well-established in many preclinical studies, its role in humans has begun to gain wider acceptance. In contrast, the role of flavonoids is mostly controversial in clinical trials, due to the diversity of the various classes of these compounds, and possibly due to the purity and the selected doses of the compounds. However, current preclinical and clinical studies warrant further detailed studies of these promising immunomodulatory compounds.

Lack of interference with immunogenicity of a chimeric alphavirus replicon particle-based influenza vaccine by preexisting antivector immunity.

Antivector immunity has been recognized as a potential caveat of using virus-based vaccines. In the present study, an alphavirus-based replicon particle vaccine platform, which has demonstrated robust immunogenicity in animal models, was tested for effects of antivector immunity on immunogenicity against hemagglutinin of influenza virus as a target antigen and efficacy for protection against lethal challenge with the virus. Chimeric alphavirus-based replicon particles, comprising Venezuelan equine encephalitis virus nonstructural and Sindbis virus structural components, induced efficient protective antibody responses, which were not adversely influenced after multiple immunizations with the same vector expressing various antigens.

Use of inactivated Escherichia coli enterotoxins to enhance respiratory mucosal adjuvanticity during vaccination in swine.

In order to augment responses to respiratory vaccines in swine, various adjuvants were intranasally coadministered with a foot-and-mouth disease virus (FMDV) antigen to pigs. Detoxified Escherichia coli enterotoxins LTK63 and LTR72 enhanced antigen-specific mucosal and systemic immunity, demonstrating their efficacy as adjuvants for nonreplicating antigens upon intranasal immunization in swine.

Immunomodulatory properties of vitamins, flavonoids and plant oils and their potential as vaccine adjuvants and delivery systems.

INTRODUCTION: During the past century, vaccinologists have attempted to mimic pathogens in their immune-enhancing capacity. This led to the development of life-saving vaccines based on live attenuated viruses, bacteria and toxoids. Hence, intense research in vaccine adjuvant discovery has focused on toll like receptors, mutant toxins and viral and bacterial vectors. Nutritive components such as vitamins and select polyphenols also possess immunomodulating properties without the potential toxic and adverse side effects of agents that mimic danger signals. AREAS COVERED: This review pertains to immunomodulatory properties of nutritive components, that is vitamins A, C, D, E, flavonoids and plant oils, as potential vaccine adjuvants and delivery systems, covering Pubmed publication searches from 1980 through 2011. EXPERT OPINION: This relatively unexplored field of the potential of nutritive components as vaccine adjuvants holds great promise to promote the development of effective and above all safe vaccines. Hence the future focus should be placed on enhancing their efficacy, mainly through novel approaches in designing structural derivatives, formulations, delivery systems and routes of administration. As safety has been the major issue in development of novel vaccines, this new approach will probably result in new discoveries in designing safe and effective vaccines.

Maintenance of long-term immunological memory by Ig+CD45R+ non-plasma B cells following mucosal immunizations.

To determine whether long-term immunological B cell memory following mucosal vaccinations is maintained by terminally differentiated Ig-CD45R- plasma cells or Ig+CD45R+ B cells, we immunized mice orally with the non-toxic B subunit of cholera toxin (CTB) as a carrier protein haptenated with FITC (CTB-FITC) plus CT adjuvant. We found that the adoptive transfer of Ig+CD45R+ but not the Ig-CD45R- cells, resulted in higher numbers of FITC-specific IgA-secreting cells in the intestine as well as higher anti-FITC serum IgA titers, suggesting that long term B cell immunological memory following oral vaccinations preferentially resided within the Ig+CD45R+ B cell population.

A novel retinoic acid, catechin hydrate and mustard oil-based emulsion for enhanced cytokine and antibody responses against multiple strains of HIV-1 following mucosal and systemic vaccinations.

Non-replicating protein- or DNA-based antigens generally require immune-enhancing adjuvants and delivery systems. It has been particularly difficult to raise antibodies against gp120 of HIV-1, which constitutes an important approach in HIV vaccine design. While almost all effort in adjuvant research has focused on mimicking the pathogens and the danger signals they engender in the host, relatively little effort has been spent on nutritive approaches. In this study, a new nutritive immune-enhancing delivery system (NIDS) composed of vitamin A, a polyphenol-flavonoid, catechin hydrate, and mustard oil was tested for its adjuvant effect in immune responses against the gp120 protein of HIV-1(CN54). Following a combination of two mucosal and two systemic vaccinations of mice, we found significant enhancement of both local and systemic antibodies as well as cytokine responses. These data have important implications for vaccine and adjuvant design against HIV-1 and other pathogens.

Mucosal HIV transmission and vaccination strategies through oral compared with vaginal and rectal routes.

IMPORTANCE OF THE FIELD: There are currently over thirty million people infected with HIV and there are no vaccines available to prevent HIV infections or disease. The genitourinary, rectal and oral mucosa are the mucosal HIV transmission routes. An effective vaccine that can induce both systemic and local mucosal immunity is generally accepted as a major means of protection against mucosal HIV transmission and AIDS. WHAT THE READER WILL GAIN: Structure and cells that comprise the oral, vaginal and rectal mucosa pertaining to HIV transmission and vaccination strategies through each mucosal route to prevent mucosal and systemic infection will be discussed. AREAS COVERED IN THIS REVIEW: Covering publications from 1980s through 2010, mucosal transmission of HIV and current and previous approaches to vaccinations are discussed. TAKE HOME MESSAGE: Although oral transmission of HIV is far less common than vaginal and rectal transmissions, infections through this route do occur through oral sex as well as vertically from mother to child. Mucosal vaccination strategies against oral and other mucosal HIV transmissions are under intensive research but the lack of consensus on immune correlates of protection and lack of safe and effective mucosal adjuvants and delivery systems hamper progress towards a licensed vaccine.

Antibody-mediated protection against mucosal simian-human immunodeficiency virus challenge of macaques immunized with alphavirus replicon particles and boosted with trimeric envelope glycoprotein in MF59 adjuvant.

We have previously shown that rhesus macaques were partially protected against high-dose intravenous challenge with simian-human immunodeficiency virus SHIV(SF162P4) following sequential immunization with alphavirus replicon particles (VRP) of a chimeric recombinant VEE/SIN alphavirus (derived from Venezuelan equine encephalitis virus [VEE] and the Sindbis virus [SIN]) encoding human immunodeficiency virus type 1 HIV-1(SF162) gp140DeltaV2 envelope (Env) and trimeric Env protein in MF59 adjuvant (R. Xu, I. K. Srivastava, C. E. Greer, I. Zarkikh, Z. Kraft, L. Kuller, J. M. Polo, S. W. Barnett, and L. Stamatatos, AIDS Res. Hum. Retroviruses 22:1022-1030, 2006). The protection did not require T-cell immune responses directed toward simian immunodeficiency virus (SIV) Gag. We extend those findings here to demonstrate antibody-mediated protection against mucosal challenge in macaques using prime-boost regimens incorporating both intramuscular and mucosal routes of delivery. The macaques in the vaccination groups were primed with VRP and then boosted with Env protein in MF59 adjuvant, or they were given VRP intramuscular immunizations alone and then challenged with SHIV(SF162P4) (intrarectal challenge). The results demonstrated that these vaccines were able to effectively protect the macaques to different degrees against subsequent mucosal SHIV challenge, but most noteworthy, all macaques that received the intramuscular VRP prime plus Env protein boost were completely protected. A statistically significant association was observed between the titer of virus neutralizing and binding antibodies as well as the avidity of anti-Env antibodies measured prechallenge and protection from infection. These results highlight the merit of the alphavirus replicon vector prime plus Env protein boost vaccine approach for the induction of protective antibody responses and are of particular relevance to advancing our understanding of the potential correlates of immune protection against HIV infection at a relevant mucosal portal of entry.

Prolonged protection against Intranasal challenge with influenza virus following systemic immunization or combinations of mucosal and systemic immunizations with a heat-labile toxin mutant.

Seasonal influenza virus infections cause considerable morbidity and mortality in the world, and there is a serious threat of a pandemic influenza with the potential to cause millions of deaths. Therefore, practical influenza vaccines and vaccination strategies that can confer protection against intranasal infection with influenza viruses are needed. In this study, we demonstrate that using LTK63, a nontoxic mutant of the heat-labile toxin from Escherichia coli, as an adjuvant for both mucosal and systemic immunizations, systemic (intramuscular) immunization or combinations of mucosal (intranasal) and intramuscular immunizations protected mice against intranasal challenge with a lethal dose of live influenza virus at 3.5 months after the second immunization.

Induction of broad CD4+ and CD8+ T-cell responses and cross-neutralizing antibodies against hepatitis C virus by vaccination with Th1-adjuvanted polypeptides followed by defective alphaviral particles expressing envelope glycoproteins gpE1 and gpE2 and nonstructural proteins 3, 4, and 5.

Broad, multispecific CD4(+) and CD8(+) T-cell responses to the hepatitis C virus (HCV), as well as virus-cross-neutralizing antibodies, are associated with recovery from acute infection and may also be associated in chronic HCV patients with a favorable response to antiviral treatment. In order to recapitulate all of these responses in an ideal vaccine regimen, we have explored the use of recombinant HCV polypeptides combined with various Th1-type adjuvants and replication-defective alphaviral particles encoding HCV proteins in various prime/boost modalities in BALB/c mice. Defective chimeric alphaviral particles derived from the Sindbis and Venezuelan equine encephalitis viruses encoding either the HCV envelope glycoprotein gpE1/gpE2 heterodimer (E1E2) or nonstructural proteins 3, 4, and 5 (NS345) elicited strong CD8(+) T-cell responses but low CD4(+) T helper responses to these HCV gene products. In contrast, recombinant E1E2 glycoproteins adjuvanted with MF59 containing a CpG oligonucleotide elicited strong CD4(+) T helper responses but no CD8(+) T-cell responses. A recombinant NS345 polyprotein also stimulated strong CD4(+) T helper responses but no CD8(+) T-cell responses when adjuvanted with Iscomatrix containing CpG. Optimal elicitation of broad CD4(+) and CD8(+) T-cell responses to E1E2 and NS345 was obtained by first priming with Th1-adjuvanted proteins and then boosting with chimeric, defective alphaviruses expressing these HCV genes. In addition, this prime/boost regimen resulted in the induction of anti-E1E2 antibodies capable of cross-neutralizing heterologous HCV isolates in vitro. This vaccine formulation and regimen may therefore be optimal in humans for protection against this highly heterogeneous global pathogen.

A comparison of anionic nanoparticles and microparticles as vaccine delivery systems.

The objective of this work was to conduct an in vivo comparison of nanoparticles and microparticles as vaccine delivery systems. Poly (lactide-co-glycolide) (PLG) polymers were used to create nanoparticles size 110 nm and microparticles of size 800-900 nm. Protein antigens were then adsorbed to these particles. The efficacy of these delivery systems was tested with two protein antigens. A recombinant antigen from Neisseria meningitides type B (MenB) was administered intramuscularly (i.m.) or intraperitonealy (i.p.). An antigen from HIV-1, env glycoprotein gp140 was administered intranasally (i.n.) followed by an i.m. boost. From three studies, there were no differences between the nanoparticles and micro-particles formulations. Both particles led to comparable immune responses in mice. The immune responses for MenB (serum bactericidal activity and antibody titers) were equivalent to the control of aluminum hydroxide. For the gp140, the LTK63 was necessary for high titers. Both nanoparticles and microparticles are promising delivery systems.

Dynamics of acute and memory mucosal and systemic immune responses against HIV-1 envelope following immunizations through single or combinations of mucosal and systemic routes.

In this study, immunizations at 2 weeks vs. 6 weeks intervals, with an HIV-1 envelope protein in adjuvants, through intra-nasal (IN), intra-muscular (IM), IN followed by IM (IN/IM) and IM/IN, were compared for induction of mucosal and systemic immune responses. IN/IM immunizations at 2, but not at 6, week intervals induced the highest mucosal and systemic immune responses compared to other immunization routes. Following a resting memory phase, IN boosting of IN/IM-immunized mice, compared to IM-boosting of IM-immunized mice, induced increased IgA responses. Thus, depending on the immunization intervals, IN/IM may be more effective than IM immunizations for short- and long-term immunity.

Protection of macaques against vaginal SHIV challenge by systemic or mucosal and systemic vaccinations with HIV-envelope.

BACKGROUND: Worldwide, the majority of human immunodeficiency virus (HIV) infections occur by heterosexual transmission. Thus, the development of a vaccine that can prevent intravaginal HIV infection is an important goal of AIDS vaccine research. OBJECTIVES: To determine which single or combination of systemic and mucosal routes of immunizations of female rhesus macaques with an HIV-1 SF162 envelope protein vaccine induced protection against intravaginal challenge with SHIV. DESIGN: Female rhesus macaques were immunized with an HIV-1 SF162 envelope protein vaccine administered systemically (intramuscularly), or mucosally (intranasally), or as a sequential combination of both routes. The macaques were then challenged intravaginally with SHIV SF162P4, expressing an envelope that is closely matched (homologous) to the vaccine. RESULTS: Macaques receiving intramuscular immunizations, alone or in combination with intranasal immunizations, were protected from infection, with no detectable plasma viral RNA, provirus, or seroconversion to nonvaccine viral proteins, and better preservation of intestinal CD4+ T cells. Serum neutralizing antibodies against the challenge virus appeared to correlate with protection. CONCLUSIONS: The results of this study demonstrate that, in the nonhuman primate model, it is possible for vaccine-elicited immune responses to prevent infection after intravaginal administration of virus.

Beta7-integrin-independent enhancement of mucosal and systemic anti-HIV antibody responses following combined mucosal and systemic gene delivery.

Vaccination strategies that can block or limit heterosexual human immunodeficiency virus (HIV) transmissions to local and systemic tissues are the goal of much research effort. Herein, in a mouse model, we aimed to determine whether the enhancement of antibody responses through mucosal and systemic immunizations, previously observed with protein-based vaccines, applies to immunizations with DNA- or RNA-based vectors. Intranasal (i.n.) followed by intramuscular (i.m.) immunizations (i.n./i.m.) with polylactide-coglycolide (PLG)-DNA microparticles encoding HIV-gag (PLG-DNA-gag) significantly enhanced serum antibody responses, compared with i.m., i.n. or i.m. followed by i.n. (i.m./i.n.) immunizations. Moreover, while i.n./i.m., i.n. or i.m./i.n. immunizations with PLG-DNA-gag resulted in genital tract antibody responses, i.m. immunizations alone failed to do so. Importantly, beta7-deficient mice developed local and systemic antibody responses following i.n./i.m. immunization, or immunization via any other route, similar to those of wild-type mice. To compare the DNA with an RNA delivery system, immunizations were performed with VEE/SIN-gag replicon particles, composed of Venezuelan equine encephalitis virus (VEE) replicon RNA and Sindbis surface structure (SIN). i.n./i.m., compared with any other immunizations, i.n./i.m. immunization with VEE/SIN-gag resulted in enhanced genital tract but not serum antibody responses. These data show for the first time that mucosal followed by systemic immunizations with gene delivery systems enhance B-cell responses independent of the mucosal homing receptors alpha4beta7 and alphaEbeta7.