WHO monographs on selected medicinal plants (detail) VOLUME 1, World Health Organization Geneva 1999 http://whqlibdoc.who.int/publications/1999/9241545178.pdf WHO monographs on selected medicinal plants Definition Herba Echinaceae Purpureae consists of the fresh or dried aerial parts of Echinacea purpurea (L.) Moench harvested in full bloom (Asteraceae) ( 1 ). Synonyms Brau
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Toll-Like Receptor Signaling: Emerging Opportunities in Human Diseases
Rabindra N. Bhattacharjee2 and Shizuo Akira*,1,2 1Department of Host Defense, Research Institute for Microbial Diseases, Osaka University Japan 2ERATO, Japan Science and Technology Agency, 3-1 Yamada, Suita city, Osaka, 565-0871, Osaka, Japan Abstract: The family of Toll-like receptors (TLR1-TLR11) provides host defense in mammals by inducing pro-
inflammatory innate immune response upon recognition of conserved structural component in pathogens. TLR
mediated activation of signaling pathways that induce the expression of proinflammatory molecules is one of
the well-studied but ever expanding fields of immunology. As a result, a wealth of information has been
obtained which includes the identification of specific ligands of individual TLR, elucidation of their
downstream signaling pathways, function of different adaptor proteins, activation of protein kinases and
transcription factors that transcribe the genes for inflammatory molecules. TLRs not only sense microbial
invasion but also can be activated by endogenous molecules as well as low molecular weight synthetic
compounds. Given the role of innate immune machinery to provoke inflammation in host, TLRs signaling may
be involved in many acute and chronic inflammatory processes in sterile and post-infection conditions such
as, atherosclerosis, leprosy, inflammatory bowel syndrome (IBD), lung airway hyperactivity in allergic asthma,
and in sepsis. By the same token, TLRs can also be associated with autoimmune diseases such as systemic
lupus erythematosus (SLE) or other immune unresponsive diseases like cancer. In addition, synthetic organic
compounds which enhance the function of TLRs can also be useful as potential adjutants to improve
conventional vaccination strategy. Here we summarize the recent development on possible modulation of the
TLR signaling pathway for therapeutic solution of multiple immune-related diseases.
Keywords: Toll like receptors, agonist-antagonist, inflammation, allergic asthma, cancer, atherosclerosis and vaccine.
microorganisms and initiate downstream signal to producereactive oxygen species (ROS), inflammatory cytokines, The early or innate response to infections by pathogens interferon and chemokines as protective measures to defend has long been considered as a non specific mean of host hosts. Up-regulation of these molecules can be a prerequisite defense. In 1996, discovery of a key Drosophila protein, for initiating and triggering a signaling cascade in cells for Toll, was shown to be required not only in embryonic the development of antigen-specific adaptive immune development for flies but also to provoke an effective immune response against the fungus Aspergillus fumigatus Thus it is plausible, given the role of TLRs in the . Since the discovery, significant advances in our induction of such strong inflammation, that improper understanding of the function of Toll proteins have evolved regulations of signaling pathway mediated by these proteins with the evidence that it is the sensing molecule in innate may also be involved in multiple inflammatory diseases immunity. Eleven Toll-like receptors (TLR1-TLR11) in such as sepsis and cancer. Besides TLR’s ability to human and 12 in mice have been identified to date. TLRs recognize PAMPs, they play an essential role in non- are type I transmembrane proteins expressed primarily in infectious sterile inflammation and are well enough to detect immune cells responsible for the first line of defense endogenous ligands from the damaged cells such as β- including macrophage, dendritic cells, mucosal epithelial defensins, and oxidized lipids . There are speculation that cells, neutrophils and dermal endothelial cells . This class TLRs may also be responded by protein molecules modified of receptors (also called pattern recognition receptors; PRR) by oxidation, and nitration . These results demonstrate is capable of sensing organisms ranging from protozoa to that TLRs are not restricted to microbial PAMPs but bacteria, fungi to viruses upon detection of the pathogen recognize wide variety of ligands. More importantly, low associated molecular patterns (PAMPs) and the signal molecular weight synthetic molecules with no obvious transduction pathway activated by them is of great structural similarity to PAMPs can successfully activate importance in evolution because it is highly conserved in signal transduction mediated by TLRs. It raises the plants, drosophila, nematode, avian and mammals [3, 4]. In possibility that blocking TLR pathway in acute-chronic vertebrates, the main function of innate immune machinery inflammation or enhancing the signal in immune is to confer the presence of PAMPs on invading unresponsive diseases might offer new ways of therapeuticintervention. In this review, we discuss the implication ofTLRs signaling in the development and progression ofhuman diseases, current use of TLR agonists and antagonists *Address correspondence to this author at the ERATO, Japan Science andTechnology Agency, 3-1 Yamada, Suita city, Osaka, 565-0871, Osaka, in experimental medicine, and future optimism of designing Japan; Tel: +81-66-879-8303; Fax: +81-66-879-8305; E-mail:
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2005 Bentham Science Publishers Ltd.
82 Current Immunology Reviews, 2005, Vol. 1, No. 1
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TLRS , LIGANDS, AND SIGNALING PATHWAYS
TLRS AND HUMAN DISEASES
Although ten (TLR1-TLR10) members of TLR protein The knowledge of innate immunity and mechanistic family have been found in human, their mechanism of details of TLRs obtained to date, most of them have been function including identification of exogenous and emerged from gene targeted mice. Although mice and endogenous ligands have emerged from gene knock out humans have many overlapping TLR events, humans may mice. The specificity of different TLRs and microbial not require a specific TLR or a particular TLR such as ligands has been elaborately demonstrated in many previous TLR11 is simply non-functional (due to polymorphism) in reviews [6, 7]. TLR2 propagates signal by binding to humans. TLR11 which detects urophathogenic E.coli microbial di- or triacylated lipoprotein, and yeast zymosan, (UPEC) doesn’t express in humans because it carries a TLR3 to double stranded viral RNA, TLR4 to endoxin common stop codon in its open reading frame (in NCBI LPS, TLR5 to bacterial flagellin , TLR7-8 to single human genome sequence, human 293 and jurkat cells). This stranded RNA, synthetic imidazoquinolines, and loxoribine may make humans prone to frequent urinary tract infection and TLR9 to CpG motifs of the bacterial DNA. An . Studying improper TLR signaling in human diseases is additional protein termed, MD-2, has been shown to be more complicated than inbred mice because of unique indispensable for signaling via TLR4. TLRs and interleukin genetic background of an individual and association of 1 receptors ( IL-1Rs) share significant similarity in their complex environmental factors. Nevertheless, studies of cytoplasmic domain (called Toll-IL-1R or TIR domain) and people with specific mutation in TLR or TLR-related genes transduce signal upon interacting with common adaptor established the fact that TLR signaling ultimately affect the protein, MyD88 (myeloid differentiation factor) that also development and progression of several human diseases. For contains a TIR domain. One or more of the three other TIR example, decreased airway sensitivity to inhaled domain containing adaptor proteins including MAL/TIRAP lipopolysaccharide (LPS) in humans was associated with a (MyD88-adaptor-like/TIR-associated protein), TRIF (Toll- TLR4 polymorphism of an amino acid substitution at the receptor-associated activator of interferon also known as position of 299 (Aspartic acid299glycine or D299G) [10, 11] TICAM-1), and TRAM (toll-receptor-associated molecule, suggesting that TLR4 is critical in the response to Gram- also known as TICAM-2) are required for TLRs activation negative bacteria. Although its mechanism of action is still by PAMPs. Several other adaptors such as SARM (sterile unknown, TLR4 protein and mRNA are upregulated in alpha and HEAT/Armadillo motif protein), and TOLLIP plaques of atherosclerotic lesions . But people carrying
(Toll-interacting protein) have recently been identified but the similar polymorphism in TLR4 have shown with lower their function in TLR signaling remain elusive. Interaction concentration of proinflammmatory cytokines and reduced between TLRs and adaptors are known as TIR-TIR progression of artery arthrosclerosis . Currently, there is interaction which activates a serine/threonine kinase family growing interest in the field to study whether TLR4 IRAK (IL-1R-associated kinase, IRAK1, IRAK2, IRAK4, polymorphism including D299G may have any effect in IRAK-M) and subsequently leads to recruitment of tumor diseases like asthma, atopy and sepsis which can also be necrosis factor (TNF) – receptor-associated factor 6 occurred by endotoxin exposure from Gram- negative (TRAF6). IRAK1/IRAK4 and TRAF6 then dissociate from infections . It has been reported that people who are the complex and become part of a new complex composed of asthmatic with this mutation, when further exposed to LPS- growth factor-β-activated kinase 1 (TAK1), TAB (TAK1 trapped-house dust, do not exacerbate the condition of binding proteins) 1 and TAB2. This formation is necessary bronco-hyper reactivity (BHR) or airway hyper-reactivity to activate TAK1 which in turn activates IκB kinase kinase (AHR) . Similarly, mutations found in human TLR2 (IKK) complex containing IKKγ or NEMO (NF-κB (R753Q), TLR2 (R677W), IRAK4 (Stop codon at 287), and essential modulator). IKK-mediated Phosphorylation of IκB IκB (missense Serine 32) may predispose people to leads to its ubiquitination and degradation thereby allowing staphylococcal infection , tuberculosis , pneumonia NF-κB to be translocated into the nucleus . Another  and immunodeficiency  respectively. It is now major event occurred in TLR signaling is the induction of obvious that TLR signaling pathway and genesis of immune interferon regulatory factor IRF3 and IRF7. During viral diseases are intimately linked. Therefore, modulation of this infection, transcription factor IRF3 is phosphorylated upon pathway might have immense possibilities in therapeutic TLR3 and TLR4 signaling (Myd88-independent but TRIF- dependent). Phosphorylated IRF3 translocates in the nucleusas homodimer and activates promoters containing IRF3binding sites. Recently IRF7 has been found in a complex IMMUNOSTIMULATION IN CANCER
with MyD88 and TRAF6 and induction of IFNα throughTLR7, TLR8 and TLR9 mediated pathways has been shown A ground breaking study in our laboratory have to be regulated by IRF7 . Activated NF-κB further identified a Toll-like receptor (TLR9) that recognizes activates transcription of many pro-inflammatory molecules unmethylated CpG oligodinucleotides (bacterial CpG ODN) such as interleukin (IL-1, IL-6, IL-8, and IL-12), tumor and activates mammalian immune cells . Now that CpG necrosis factor (TNF), cyclooxygenase-2 (COX-2) and nitric molecules can specifically interact with TLR9 to enhance oxide synthase (iNOS) (Fig. 1). IRF3 controls the
immunogenicity, use of various synthetic organic molecules production of type 1 interferon (IFN) β and IFNβ is in modulation of TLR signaling creates a momentum in necessary for the expression of IRF7 to enhance induction of therapeutic medicine. Without knowing the mechanism of other interferon (IFNα and IFNβ). All of these molecules are action, CpG has long been known as a strong adjuvant and essential for successful removal of pathogens and alert host its use in human hepatitis vaccine has greatly improved immunogenicity . Biological effects mediated by CpG Toll-Like Receptor Signaling
Current Immunology Reviews, 2005, Vol. 1, No. 1 83
Fig. (1). Signaling pathways induced by TLRs and their microbial and synthetic ligands. Signal transduction is initiated by both
transmembrane and intracellular TLRs (TLR7, TLR8 and TLR8). Most of the TLRs are functional as homodimer but TLR1 and TLR6 do
form heterodimer with TLR2 to discriminate differences between triacyl and diacyl lipopeptides, respectively. Activation of TLRs
triggers interaction with the adaptor molecule MyD88. Other adaptor molecules such as TIRAP, TRIF and TRAM are also involved.
TLR2 recognizes microbial lipopeptides (LP), lipoteic acid (LTA) and yeast zymosan. TLR4 is the receptor for LPS. TLR5 is essential
in flagellin recognition, whereas TLR9 detects CpG DNA. Mouse TLR7 and human TLR8 are specific for single-stranded RNA as well
as synthetic molecules imiquimod (R-837) and resiquimod (R-848). The MyD88 dependent pathway finally leads to the translocation
of NF-kB into nucleus which regulates the expression of many genes related to proinflammatory mediators. TLR3 and TLR4 can also
induce a TRIF-dependent but MyD88-independent pathway leading to the activation of IRF-3 and then IFN-β. Several synthetic
TLR4, TLR7, TLR8 and TLR9 agonists and antagonists
such as CpG7909, loxorobine, MLA, MPL, RC-529, Poly I:C, compound 4a, chloroquine etc. have been shown to stimulate or abrogate TLR signaling and should be useful for treating various immune-relatedhuman diseases. Compound 4a mimics adaptor protein MyD88 and prevents TIR-TIR interaction.
now appeared to be far more comprehensive. It directly tumor cells from metastatic Lewis lung carcinoma (LLC), activates dendritic cells to secrete IL6, IL12, GM-CSF survival time were extended significantly upon CPG 7909 (granulocyte-macrophage colony stimulating factor), treatment. The survival time can be further enhanced when chemokine, TNF, and IFNα. These cytokines then combination of paclitaxel (well-known chemotherapeutic stimulates natural killer (NK) cells to secret IFNγ . CpG agent) and CPG 7909 were used . Patients with promotes proliferation of B cells that ultimately secrete IL6, advanced form of NSCLC (stage III/IV) and beyond surgical IL12 and polyclonal IgGs. A clear predominance of IL12 and cure will be receiving this treatment for clinical efficacy and IFNγ with almost no secretion of TH-2 type cytokines indicates that CpG induces much appreciated T-helper 1 Signaling from TLR7 and TLR8 might also have (TH1) type adaptive immune response (Fig. 2) . Because
potential significance as a target for cancer therapy. Because of CpGs immunogenic and or immunostimulatory in part, two small synthetic antiviral compounds imiquimod properties, one can easily speculate that it may have anti- (R-837) and resiquimod (R-848) which are well known tumor activities too. Therefore, synthetic derivatives of CpG have gained wide popularity among drug designers as a imidazoquinolines also work as ligands for these receptors.
therapeutic target to treat allergy, asthma and cancer.
Imiquimod and resiquimod direct their signal upon binding Scientists at Coley’s TLRTM Therapeutic synthesized a wide to endosomal (intracellular) TLR7 and 8. Using MyD88 range of molecules based on alteration in the number of the deficient mice, it has been demonstrated that signaling sequences of CpG. One of their leading compound, occurs through the TLR-MyD88 pathway. Much of the ProMuneTM (CPG 7909), is currently in phase III clinical significant activity has been attributed to the ability of these trial to treat advanced non small cell lung cancer (NSCLC), compounds to induce production of IFNs. Increased IFNα malignant melanoma, and cutaneous T cell lymphoma and γ are observed in human blood cells treated with (CTCL). Experimental data indicates that mice harboring 84 Current Immunology Reviews, 2005, Vol. 1, No. 1
Bhattacharjee and Akira
Fig. (2) Mechanism of TLR9 signaling mediated by CpG oligonucleotides. A diverse effect including a TH-1 type immune
response, natural killer cell activity, dendritic cell activation, cytokine, and interferon-γ production have been found for CpG.
CpG initiates both an innate and adaptive immune response by generationg cytotoxic T cell, and antigen specific antibody
production. IL, interleukin; TNF, tumor necrosis factor; MHC, major histocompatibility complex.
imiquimod in vitro . Upregulation of IFNs subsequently
proven safe to use in human. From the results of clinical induces the production of other cytokines such as IL1, IL6 observations, there is a growing optimism that development and IL8 in various cell types . Imidazoquinolines have of new agonists of TLR7, 8 and 9 could be beneficial to shown to be effective against viral infections such as human papillomavirus, herpes simplex virus and have also beensuccessfully used for the treatment of intra-anal condyloma,genital warts, and Mollusca contagiosa . Interestingly, ALLERGY AND ASTHMA
topical application of imiquimod on cancerous lesions fromsquamous cell carcinoma greatly improved the condition and Allergic diseases including asthma have reached epidemic now it is being used in human treatment . Both in vitro proportions in the world, affecting approximately 150-200 and in vivo studies indicate that imiquimod (not resiquimod) million people worldwide in all ages and races (World induced apoptosis in melanoma cells in a tumor selective Health Organization report, 2004). The number of asthma manner . Moreover, topical imiquimod has been patients is growing by 50% every decade and causes 200,000 evaluated in dozens of other cancers including superficial deaths a year. Asthma is a chronic, inflammatory lung basal cell cancer, Bowen’s disease and melanoma . Many disease associated with bronchial hyper-reactivity, and of them are in phase II/ III trials and 50% to complete airway inflammation characterized by recurrent breathing clinical responses have been reported elsewhere. Meanwhile problem. Among many known triggers for asthma, allergens another compound, loxoribine (7-allyl-7, 8-dihydro-8-oxo- contain endotoxin (LPS) such as mold, flu virus, and house guanosine), a MyD88 dependent TLR7 agonist, is now dust have been studied well. One of the predominant under clinical trial (Phase I) for the evaluation of its anti- mechanisms in asthma is the generation of allergen-specific tumor activities in patients with advanced cancer .
TH-2 type (T helper cells type 2) response that produces Loxoribine showed limited toxicity to mammalian cells and interleukins necessary of the development of TH-2 cells(IL4), regulation of immunoglobulin E (IgE) production Toll-Like Receptor Signaling
Current Immunology Reviews, 2005, Vol. 1, No. 1 85
(IL13) and accumulation of Eosinophils (IL5) [31, 32].
(rhinovirus and coronavirus) are the most frequent triggers of Activated mast cells upon IgE cross linking and TH-2 cells asthma in both children and adults. As we already know that further stimulate the production of histamines, and synthetic imidazoquinolines (imiquimod and resiquimod) chemokines which then lead to airway obstruction, hyper- are antiviral in function which can induce IFNγ as well.
responsiveness, and mucus overproduction in asthma Considering the role of viral infection in disease patients . Importantly, TH-2-like T cells failed to exacerbations of asthma, their use might have considerable produce IL2 or IFN γ which are thought to be involved in impact in improving the quality of life of asthma patients.
protective mechanism in allergic asthma. There are evidences Not surprisingly, these drugs are already in use in the clinic that children who encounter less microbial infections for to treat human warts caused by papilloma virus and are being brought up in an extremely hygienic condition in the proven safe. Furthermore, a single application of resiquimod developed world, produce decreased TH-1 type response and (R-848) in animal prevented allergen-induced development thereby increased TH-2 type inflammation. One possible of airway hyperreactivity and inflammation in vivo . A explanation that TH-1 mediated immune response produces switch of cytokine expression from a prevalent TH-2 type IFNγ which inhibits both IgE synthesis and eosinophilia towards a predominant TH-1 type has also been reported in [34, 35]. Corticosteroids and bronchodilators are the only these mice. If this is the case in human, TLR7 and 8 drugs available now to treat asthma. Although the drugs are agonists can work like a double- edged-sword in asthma effective in relieving the symptoms temporarily, they are battle by redirecting disease mechanism as well as protecting unable to shift the existing TH-2-cell-type response in from disease exacerbations due to viral attack.
sensitized individuals. The demand for a more specificasthma treatment has expected to grow worldwide.
Numerous preclinical studies based on modulation of the ATHEROSCLEROSIS AND TLR4
TLR signaling opened an important area of innateimmunotherapy to be studied for new drugs for asthma. The Atherosclerosis is now considered to be a chronic potential drug against asthmatic inflammation would have to inflammatory disease. A numerous studies indicate that be able to reverse one strong manifestation of asthma; T TLR4 and MyD88-mediated signaling play an essential role like pattern of cytokine production (IL4, IL5, IL13 etc.) to in the initiation and progression of atherosclerosis.
Compared to unaffected vessels, both TLR4 mRNA and H-1 like response (IL12, IFNγ etc ). Not unexpectedly, CpG ODN in mice model induces the production of T proteins are highly expressed in atherosclerotic lesions .
Oxidized LDL (low density lipoprotein) is a known trigger episodes of bronchial hyperreactivity and airway eosinophilia for atherogenesis. Observation that macrophages stimulated by LDL induce an upregulation of TLR4 protein expression critical for the development of airway hypereractivity .
in vitro, suggests a clear role of TLR4 in lipid-mediated pro- Subsequent study in a murine model confirms that CpG inflammatory signaling . In an atherosclerotic mouse ODN is capable to reverse allergic inflammation by model treated with LPS, a notable increase in plaque area inhibiting IL13 and goblet cell hyperplasia . More provides a clear evidence of TLR4 participation in plaque recently, Spiegelberg HL et al.  made an allergen- CpG formation . Genetic polymorphism study in human has ODN conjugate by covalent binding (also known as also shown that a mutation in TLR4 ( Asp299Gly) is alergoid) and applied this novel approach in mouse model of associated with less thickness in the carotid artery and allergy. Compare with CpG-ODN alone, this conjugate were reduced risk for carotid artery atherosclerosis . Very recently, another group used apolipoprotein E –deficient mouse (known model for atherosclerosis) to see the effect of asthma have also been reversed considerably. Phase I/II trials TLR signaling in the disease progression .
in human ragweed patients showed that the ragweed-CpG Apolipoprotein E –deficient mice that also lacked TLR4 or ODN conjugates are safe and well tolerated. Patients were its downstream adapter molecule MyD88 exhibited decreased less allergenic and no significant increase in IgE and aortic atherosclerosis accompanied by reduced plaque lipid histamine were observed. In contrast, anti-allergen antibodies content, proinflammatory cytokines IL12, macrophage (IgG) were appeared more rapidly in patients treated with infiltration, and cyclooxygenase (COX) -2 immunore- conjugate than allergen itself. It is interesting to see that the activity. This work has provided a conclusive evidence in responses of clinical compounds in animal studies can be establishing a link between MyD88-dependent TLR4 effectively reproduced in human diseases. Keeping this in signaling and hypercholesterolemia and or atherosclerosis.
mind, therapeutic drug developers are synthesizing varieties Therefore, a specific antagonist for TLR4 signaling pathway of CpG –ODNs for next generation of drug discovery based may serve as a valuable tool in reducing the harmful effects on TLR mechanism. Two of such yet undisclosed structural of inflammation and other biological consequences in many compounds are now approved to use in human clinical diseases including atherosclerosis. Unfortunately no pharma - testing under a group of pharmaceutical companies.
ceutical TLR4 antagonist exists to date so far. Meanwhile anexciting piece of work has been reported on a newly While asthma itself is a significant burden to a patient, synthesized cell-penetrating-small-molecule, hydrocinnamoyl disease exacerbations in asthma (i.e., wheezing, coughing –L-valyl pyrrolidine (or compound 4a). Compound 4a pain, tightness of chest and breathe shortness) are accounted mimics the essential TLR adapter protein, MyD88 and for most of the discomfort, cost and morbidity. Although interferes with the interaction between mouse MyD88 and environmental pollutants can aggravate the asthma episode TLR/IL-1R at the TIR domain (TIR-TIR protein interaction) to some degree, viral infections contribute more than 60% thus inhibits IL-1β-induced signaling pathways . We cases of wheezing in children . Common cold viruses have already described the importance of TIR-TIR mediated 86 Current Immunology Reviews, 2005, Vol. 1, No. 1
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homotypic interaction between TLRs and MyD88 in innate targeting stimulators of the inhibitory signaling pathways of immune signal transduction. Further clinical research on TLRs might have therapeutic potential in sepsis and compound 4a is necessary to explain how the blockade of autoimmune disorders. Interestingly, one group used a TLR/IL1R signaling therapeutically affects many diseases murine model of collagen induced-arthritis to see the effect including rheumatic arthritis, sepsis and atherosclerosis. The of soluble ST2-Fc (sST-Fc) fusion proteins. Administration availability of more and more small-molecule antagonists in of sST-Fc down-regulates serum cytokine levels (IL6, IL12, near future would be certainly helpful to evaluate the clinical and TNFα) and significantly attenuated disease severity efficacy of therapeutic inhibition of TLR signaling and their . Further studies (both experimental and clinical) are required to know if sST2 can play a beneficial role inrheumatoid arthritis (RA) and sepsis.
IMMUNOSUPPRESSION IN SEPSIS
TLR AGONISTS TO IMPROVE VACCINATION
Improper or uncontrolled TLR signaling has been blamed for the pathophysiology of many diseases including sepsis, The discovery of TLRs has significantly accelerated the inflammatory bowel diseases (IBD), autoimmune syndromes advancement of our knowledge on how components of and leprosy. Sepsis is defined as systemic inflammatory pathogens propel the immune system into an activated state.
response syndrome (SIRS) and occurs from the body’s Understanding TLR pathways opens up new direction to systemic over-reaction to infection (bacterial, viral, fungal, tackle a range of challenges still exist in medical or parasitic). This over-reaction disrupts homeostasis immunology including vaccination. One of the fields that through a dysregulated cascade of inflammation, coagulation could greatly improve from this discovery is the and impaired fibrinolysis which leads to global hypoxia, development of new adjuvant for vaccine. Despite then multiple organ failure and finally death . Sepsis and tremendous advances of immunology, human adjuvant its most severe form, septic shock, kill 215, 000 people remains almost totally dependent on aluminum based every year in USA only and will continue to cause havoc compound. A good adjuvant enhances the effectiveness of worldwide. Numerous attempts including anti-inflammatory vaccine by activating antigen presenting dendritic cells, B (anti-LPS, anti-cytokine) and anticoagulant treatments in the cells and induces a TH1-like immune response. These past decade have been failed. Therapy for sepsis is limited to ultimately alert adaptive immune system against the co- anti-infectives and supportive care which do not treat the administrated antigen. The discovery that some agonists of underlying pathophysiology. Microbiological diagnosis TLR can directly activate dendritic cells, manipulation of indicates that gram-negative sepsis account for about 60% of innate immune system might lead to more effective vaccines the total cases and LPS is a potent and predominant against cancer, autoimmune disorder, and infectious mediator that induces an intense inflammatory response diseases. TLR9 ligand CpG ODN is an excellent candidate through the innate immune system. We now know that, in this game. Adding CpG ODNs to a commercial hepatitis TLR4 is the receptor for the LPS . There are reports that B (HB) vaccine ( Engerix- BR, GlaxoSmithKline biological) bacteria carrying LPS mutation do not elicit or animals resulted in a 15-fold increase of anti-HBs IgG antibodies in lacking TLR4 do not develop septic shock in response to animal compared to vaccine alone  . Consistently, LPS [49, 50]. Taken together it raises a hope that someday preliminary result from a phase-I human trial demonstrated the devastating effects of bacterial sepsis can be partially the ability of CpG DNA to enhance immunogenicity of the resolved by manipulating TLR signaling pathway. TLR4 hepatitis B vaccine by raising anti-HB antibody titer and MD-2 antagonists as anti-inflammatory agents are in the compared to vaccine alone . Interest is growing among list of potential future drug but those do not exist now.
biopharmaceutical companies to develop a new generation of Another approach worked well in murine sepsis model novel vaccine based on TLR signaling pathways. Individual which shown that, naturally occurring soluble form of company like Corixa Corporation (Seattle, USA), OM mouse TLR4 protein successfully can block LPS signaling Pharma (Geneva, Switzerland) and Coley TLR therapeutics . Surprisingly, compound 4a (as described above) has (Wellesley, MA, USA) are developing their own been found to be very effective to block intracellular TIR- investigational TLR agonists/antagonists and many of them TIR protein interaction. Therefore, for the first time, a are already in different phases of human clinical trials. There potential intracellular site for an anti-inflammatory drug are claims from corporate researchers (Coley pharmaceuticals) action has been discovered and this phenomenon can well be that at least one of their products, Vaximmune, a short tested in multiple diseases such as atherosclerosis, rheumatic synthetic oligonucleotide which can target the TLR9 uniquely found in the dendritic and B cells and elicit apowerful immune response against cancer, and infectious Another thought emerged from a rationale that if TLRs diseases. Most importantly, vaccines contain Vaximmune can evoke such strong anti-inflammatory response in adjuvant; protective antibody levels are reached with reduced infection, they must have endogenous molecules or antigen dose and boost requirements (unpublished data).
mechanism to maintain host homeostasis by a self-limitingfeed back regulation. Indeed two members of the TIR The most promising data have been obtained using a family, SIGIRR (single immunoglobulin domain IL-1R- TLR4 adjuvant MPL (CorixaTM). MPL is chemically related)  and ST2 have been found as repressors of TLR similar to the monophosphoryl lipid A (MLA) moiety of the signaling. ST2 deficiency produces increased cytokines in bacterial endotoxin LPS but lacks an acid-labile phosphoryl macrophages, and fails to develop endotoxin tolerance in group and a base-labile acyl group. Clinical data obtained mice . A recent review by Ishii  emphasizes that from 33,000 doses of MPL adjuvant to over 12,000 Toll-Like Receptor Signaling
Current Immunology Reviews, 2005, Vol. 1, No. 1 87
individual to date has proven to be a potent negative and gram-positive bacteria is unable to induce pro- immunostimulator yet apparently non-toxic in human when inflammatory reaction. However, intestinal epithelial cells administered with a variety of viral and bacterial antigen and macrophages of the IBD patients showed increased including peptides, polysaccharides and tumor cell lysates.
expression of TLR4 and TLR2 . Another major recent MPL activates TLR4 dependent release of cytokines IL1, finding which can relate the role of innate immunity in the IL12, TNFα and GM-CSF from macrophages and development of IBD is the discovery of a nucleotide-binding monocytes. It helps to recruit dendritic cells and APCs oligomerization domain (NOD) 2. NOD2 acts like a pattern (antigen presentating cell) to the local lymphoid organs, recognition receptor and recognize bacterial peptidoglycan as increases IFNγ production followed by generation of ligand. Interestingly, a frame shift mutation caused by the complement fixing antibodies, IgG2a . Most notably, insertion of cytosine (3020insC) that produces a truncated two doses of MPL containing Engerix B was enough to NOD2 protein, is associated with Crohn’s disease (a chronic reach the level of protection (anti-HB antibodies) compare to inflammatory disorder of the Gastro-intestinal tract) .
current three doses of Engerix B. A series of related Whether or not NOD2 receptors do function independently synthetic compounds called aminoalkyl glucosaminide 4 from the TLRs is not resolved. More information requires phosaphates (AGPs) which includes RC-529 and RC-524 how targeting NOD2 and TLRs systems could pave the way (Corixa TM) also work as lipid A mimetics and act on TLR 4. RC-529 adjuvant showed seroprotection of more than Choroquine (Aralen HCL, USA brand name) is an 95% of individual after two doses of vaccination for the antiprotozoal medicine used to treat malaria worldwide.
Although the proper mechanism of action was unknown, the Few studies in mice have indicated that both imiquimod drug showed a modest efficacy in the treatment of systemic (Aldara; 3M pharmaceuticals, St Paul, MN, USA) and lupus erythematosus (SLE), rheumatoid arthritis and various resiquimod are suitable adjuvants for therapeutic DNA skin disorders [65, 66]. SLE is a chronic autoimmune disease in which the patients produce a wide range of subcutaneously at the vaccination site immediately after autoantibody specificities. A study unraveled how both particle mediated delivery of plasmid DNA, it was found to innate and adaptive immune systems were involved with increase the number and maturation of dendritic cells, IFNγ SLE. B cell plays devastating roles in SLE because it production, and enhances antigen-specific T (CD4+ and expresses the receptor for self-IgG. But B cells can only to CD8+) cell responses . A modest immune response has be activated by IgG2a-chromatin complex when dual receptor also been observed in another study using resiquimod as an signaling occurs from both B cell receptor (BCR) and TLR9 adjuvant for HIV-1 gag DNA plasmid vaccine therapy by . Consequently the drug chloroquine is found to be an intra-muscular immunization of BALB/c mice . A big another TLR9 inhibitor. This finding is unique and scale clinical trial is now underway (GlaxoSmithKline and important because two distinct pathways are involved in Wyeth) aiming TLR based effective vaccines against twenty SLE. Inhibitors specific for these pathways may lead to the development of therapies that specifically target autoreactiveB cells.
AUTOIMMUNE DISEASE AND INFECTION
Bacteria that causes leprosy, Mycobacterium leprae (M. laprae), is found to activate TLR2 and TLR2-TLR1 It has been exactly seven years since the mammalian heterodimers. TLR2 and TLR1 are strongly expressed in T- TLRs were found as the principal inducers of the innate lep lesions (localized tuberculoid form of leprosy) together immune system. The impact of their identification is among with dominant local expression of the type-1 cytokines the most important and fundamental discoveries in microbial IFNγ , IL12, IL18 and GM-CSF. Microbial lipoprotein is a pathogenesis. Today innate immunity is increasingly potent ligand for TLR2-TLR1 heterodimers and that could recognized as the central defense system because its serve as PAMP on M. laprae to be recognized by innate mechanism also empowers the adaptive immune responses.
immune machinery . Synthetic peptides corresponding Within this short period of time, remarkable advances in our to the M. laprae 19-kD and 33-kD lipoprotein were able to understanding of the identification and characterization of activate both monocyte and dendritic cells in a TLR2 dozens of molecules involved in TLRs signaling pathways dependent manner to release IL12p40. Nerve damage, a have been obtained. There is no longer any doubt that the hallmark of leprosy, has continued to happen even when TLRs are capable of sensing a wide range of microbes and pathogens are contained and TLR2 activation is strong. It is quickly produce anti-microbial chemokines, cytokines, quite possible that while TLR activation is a quick response interferon, and reactive oxygen species to protect the host to infection, subsequent non-stop inflammatory reaction due against invading pathogens. At the same time, it is practical to deregulated TLR signaling might cause tissue (nerve) to think that hyper or hypo-responsiveness of TLRs or damage in leprosy deserves further investigation.
uncontrolled and improper signaling from these receptorscould have serious consequences in hosts. Indeed, TLR Human inflammatory bowel disease (IBD) is signaling is intimately linked with many diseases. Hopes are characterized by chronic intestinal inflammation but mounting that the present knowledge on TLR biology involvement of any particular pathogen has not been allows us to design new approaches to modulate this system discovered yet. Under normal condition, intestinal epithelial in ways that favorably affect a disease condition (Table 1).
cells do not express TLRs and MD-2 . This may be Many pharmaceutical companies as well as individual because gastrointestinal microflora consisting of both gram- 88 Current Immunology Reviews, 2005, Vol. 1, No. 1
Bhattacharjee and Akira
Synthetic Agonists and Antagonists of TLRs in Various Diseases
Atherosclerosis, RA, sepsis; Blocks TIR-TIR Interaction; Intracellular antagonist Enhance quality of vaccines; Recruit APCs to lymphoid organs, increase IFN-γ Production, Generate complement fixing antibody RA, sepsis ;TLR repressor. Down regulate serum cytokines Wide range of anti-tumor activities.
Viral infection in disease exacerbations in asthma Papilloma virus infection, apoptosis inducer in melanoma Viral infections, condyloma, genital warts, Mollusca contagiosa, basal cell Suitable DNA vaccine adjuvant; Immunogenicity increased with HIV-1 gag SLE, RA, autoimmune diseases, skin disorder; TLR9 inhibitor Non small cell lung cancer, malignant melanoma cutaneous T cell lymphoma. Strong Asthma, allergy, inhibits IL13 and goblet cell Hyperplasia Pollen allergy, anti-allergen antibody production RA, Rheumatoid arthritis; TIR, Toll-interleukin-1R; SLE, systemic lupus erythematosus.
APC, antigen presenting cell.
laboratories are involved in synthesizing small molecules Ozinsky A, Underhill DM, Fontenot JD, et al. The repertoire for that can serve as TLR antagonist or agonists. Many of such pattern recognition of pathogens by the innate immune system isdefined by cooperation between toll-like receptors. Proc Natl molecules such as CpG 7909, imiquimod, resiquimod and chloroquine have proven safe to use in human. CpG based Ulevitch RJ, Therapeutic targeting the innate immune system. Nat adjutants are now under comprehensive human trials to improve vaccination strategy in many diseases. Because Akira S, Takeda K. Toll-like receptor signaling. Nature Rev innate immune response is not pathogen specific, any Takeda K, Kaisho T, Akira S. Toll-like receptors. Annu Rev intervention of TLRs pathway could have a broad spectrum of use. Toll opens up an exciting era of drug development Kawai T, Sato S, Ishii KJ, et al. Interferon-alpha induction which will continue to be flourished and significantly through Toll-like receptors involves a direct interaction of IRF7 improve the quality of human life in not-so-distant future.
with MyD88 and TRAF6. Nat Immunol 2004; 5: 1061-8.
Zhang D, Zhang G, Hayden MS, et al. A toll-like receptor thatprevents infection by uropathogenic bacteria. Science 2004; 303:1522-6.
Arbour, N.C. Lorenze, Schutte BC, et al. TLR4 mutations areassociated with endotoxin hyporesponsiveness in humans. Nat We are thankful to all members of our laboratory and ERATO office for helpful cooperation, particularly Drs. Ken Cook DN, Pisetsky DS, Schwartz DA.Toll-like receptors in the J. Ishii, Osamu Takeuchi, Taro Kawai and Satoshi Uematsu pathogenesis of human disease. Nat Immunol 2004; 5: 975-9.
Xu XH, Shah PK, Faure E, et al. Toll-like receptor-4 is expressed for technical input, Dr. Manoor P. Hande for critical reading by macrophages in murine and human lipid-rich atherosclerotic plaques and upregulated by oxidized LDL. Circulation 2001; 104:3103-3108.
Kiechl S, Lorenz E, Reind I, et al. Toll-like receptor 4 REFERENCES
polymorphisms and atherogenesis. N Engl J Med 2002; 347: 185-192.
Child NJ, Yang IA, Pulletz MCK, et al. Polymorphisms in Toll-like Lemaitre B, Nicolas E, Michaut L, Reichhart JM, Hoffmann JA.
receptor 4 and the systemic inflammatory response syndrome.
The dorsoventral regulatory gene cassette spatzle/Toll/cactus Biochem Soc Trans 2003; 31: 652–653.
controls the potent antifungal response in Drosophila adults. Cell1996; 86: 973-83.
Werner M, Topp R, Wimmer K, et al. TLR4 gene variants modifyendotoxin effects on asthma. J Allergy Clin Immunol 2003; 112: Imler JL, Hoffmann JA.Toll-like recptors in inante immunity.
Lorenz E, Mira J, Cornish K, Arbour N, Schwartz DA. Novel Hajjar AM, O'Mahony DS, Ozinsky A, et al. Cutting edge: polymorphism in the toll-like receptor 2 gene and its potential functional interactions between toll-like receptor (TLR) 2 and association with staphylococcal infection. Infect Immun 2000; 68: TLR1 or TLR6 in response to phenol-soluble modulin. J Immunol Toll-Like Receptor Signaling
Current Immunology Reviews, 2005, Vol. 1, No. 1 89
Ben-Ali M, Barbouche MR, Bousnina S, Chabbou A, Dellagi K.
Freymuth F, Vabret A, Brouard J, et al. Detection of viral, Toll-like receptor 2 Arg677Trp polymorphism is associated with Chlamydia pneumoniae and Mycoplasma pneumoniae infections susceptibility to tuberculosis in Tunisian patients. Clin Diagn Lab in exacerbations of asthma in children. J Clin Virol 1999; 13: 131- Picard C, Puel A, Bonnet M, et al. Pyogenic bacterial infections in Quarcoo D, Weixler S, Joachim RA, et al. Resiquimod, a new humans with IRAK-4 deficiency. Science 2003; 299: 2076-2079.
immune response modifier from the family of Puel A, Picard C, Ku CL, Smahi A, Casanova JL. Inherited imidazoquinolinamines, inhibits allergen-induced Th2 responses, disorders of NF-κB-mediated immunity in man. Curr Opin airway inflammation and airway hyper-reactivity in mice. Clin Hemmi H, Takeuchi O, Kawai T, et al. A Toll-like receptor Xu XH, Shah PK, Faure E, et al. Toll-like receptor-4 is expressed recognizes bacterial DNA. Nature 2000; 408: 740-5.
by macrophages in murine and human lipid-rich atherosclerotic Hartmann G, Krieg AM. Mechanism and function of a newly plaques and upregulated by oxidized LDL. Circulation 2001; 104: identified CpG DNA motif in human primary B cells. J Immunol Hollestelle SC, De Vries MR, Van Keulen JK, et al. Toll-like Finlay BB, Hancock RE. Can innate immunity be enhanced to receptor 4 is involved in outward arterial remodeling. Circulation treat microbial infections? Nat Rev Microbiol 2004; 2: 497-504.
Weeratna RD, Bourne LL, Sullivan SM, Davis HL, Krieg AM.
Michelsen KS, Wong MH, Shah PK, et al. Lack of Toll-like Combination of a new TLR9 agonist immunomodulator (CpG receptor 4 or myeloid differentiation factor 88 reduces 7909) and paclitaxel for treatment of metastatic Lewis Lung atherosclerosis and alters plaque phenotype in mice deficient in Carcinoma (LLC). ASCO annual meeting proceedings 2004; 22: apolipoprotein E. Proc Natl Acad Sci USA 2004; 101: 10679-84.
Bartfai T, Behrens MM, Gaidarova S, et al. A low molecular Weeks CE, Gibson SJ. Induction of interferon and other cytokines weight mimic of the Toll/IL-1 receptor/resistance domain inhibits by imiquimod and its hydroxylated metabolite R-842 in human IL-1 receptor-mediated responses. Proc Natl Acad Sci USA blood cells in vitro. J Interferon Res 1994; 14: 81-5.
Imbertson LM, Beaurline JM, Couture AM, et al. Cytokine Aird WC.The role of the endothelium in severe sepsis and induction in hairless mouse and rat skin after topical application of multiple organ dysfunction syndrome. Blood 2003; 101: 3765-77.
the immune response modifiers imiquimod and S-28463. J Invest Beutler B.Tlr4: central component of the sole mammalian LPS sensor. Curr Opin Immunol 2000; 12: 20-6.
Berman B. Imiquimod: a new immune response modifier for the Hoshino K, Takeuchi O, Kawai T, et al. Cutting edge: Toll-like treatment of external genital warts and other diseases in receptor 4 (TLR4)-deficient mice are hyporesponsive to dermatology. Int J Dermatol 2002; 41: 7-11.
lipopolysaccharide: evidence for TLR4 as the Lps gene product. J Hengge UR, Ruzicka T. Topical immunomodulation in dermatology: potential of toll-like receptor agonists. Dermatol Brandtzacg P, van Dcurcn M. Current concepts in the role of the host response in Neisseria meningitidis septic shock. Curr Opin Schon MP, Wienrich BG, Drewniok C, et al. Death receptor- independent apoptosis in malignant melanoma induced by the Iwami KI, Matsuguchi T, Masuda A, Kikuchi T, Musikacharoen small-molecule immune response modifier imiquimod. J Invest T, Yoshikai Y. Cutting edge: naturally occurring soluble form of mouse Toll-like receptor 4 inhibits lipopolysaccharide signaling. J Mackenzie-Wood A, Kossard S, de Launey J, Wilkinson B, Owens ML. Imiquimod 5% cream in the treatment of Bowen's Wald D, Qin J, Zhao Z, et al. SIGIRR, a negative regulator of disease. J Am Acad Dermatol 2001; 44: 462-70.
Toll-like receptor-interleukin 1 receptor signaling. Nat Immunol Agarwala SS, Kirkwood JM, Bryant J. Phase 1, randomized, double-blind trial of 7-allyl-8-oxoguanosine (loxoribine) in Brint EK, Xu D, Liu H, et al. ST2 is an inhibitor of interleukin 1 advanced cancer. Cytokines Cell Mol Ther 2000; 6: 171-6.
receptor and Toll-like receptor 4 signaling and maintains Anderson GP, Coyle AJ. TH2 and 'TH2-like' cells in allergy and endotoxin tolerance. Nat Immunol 2004; 5: 373-9.
asthma: pharmacological perspectives. Trends Pharmacol Sci Ishii KJ, Akira S. Toll-like Receptors and Sepsis. Curr Infect Dis Corry DB, Kheradmand F. Induction and regulation of the IgE Leung BP, Xu D, Culshaw S, McInnes IB, Liew FY. A novel therapy of murine collagen-induced arthritis with soluble T1/ST2.
Bousquet J, Jeffery PK, Busse WW, Johnson M, Vignola AM.
Asthma. From bronchoconstriction to airways inflammation and Krieg AM, Davis HL. Enhancing vaccines with immune remodeling. Am J Respir Crit Care Med 2000; 161: 1720-45.
stimulatory CpG DNA. Curr Opin Mol Ther 2001; 3: 15-24.
Coffman RL, Seymour BW, Lebman DA, et al. The role of helper Evans JT, Cluff CW, Johnson DA, Lacy MJ, Persing DH, T cell products in mouse B cell differentiation and isotype Baldridge JR. Enhancement of antigen-specific immunity via the regulation. Immunol Rev 1988; 102: 5-28.
TLR4 ligands MPL adjuvant and Ribi.529. Expert Rev Vaccines Iwamoto I, Nakajima H, Endo H, Yoshida S. Interferon gamma regulates antigen-induced eosinophil recruitment into the mouse Baldridge JR, Cluff CW, Evans JT, et al. Immunostimulatory airways by inhibiting the infiltration of CD4+ T cells. J Exp Med activity of aminoalkyl glucosaminide 4-phosphates (AGPs): induction of protective innate immune responses by RC-524 and Broide D, Schwarze J, Tighe H, et al. Immunostimulatory DNA RC-529. J Endotoxin Res 2002; 8: 453-8.
sequences inhibit IL-5, eosinophilic inflammation, and airway Thomsen LL, Topley P, Daly MG, Brett SJ, Tite JP. Imiquimod hyperresponsiveness in mice. J Immunol 1998; 161: 7054-62.
and resiquimod in a mouse model: adjuvants for DNA vaccination Sur S, Wild JS, Choudhury BK, Sur N, Alam R, Klinman DM.
by particle-mediated immunotherapeutic delivery. Vaccine 2004; Long term prevention of allergic lung inflammation in a mouse model of asthma by CpG oligodeoxynucleotides. J Immunol 1999; Otero M, Calarota SA, Felber B, et al. Resiquimod is a modest adjuvant for HIV-1 gag-based genetic immunization in a mouse Taube C, Duez C, Cui ZH, et al. The role of IL-13 in established allergic airway disease. J Immunol 2002; 169: 6482-9.
Krutzik SR, Ochoa MT, Sieling PA, et al. Activation and Serebrisky D, Teper AA, Huang CK, et al. CpG regulation of Toll-like receptors 2 and 1 in human leprosy. Nat oligodeoxynucleotides can reverse Th2-associated allergic airway responses and alter the B7.1/B7.2 expression in a murine Abreu MT, Vora P, Faure E, Thomas LS, Arnold ET, Arditi M.
model of asthma. J Immunol 2000; 165: 5906-12.
Decreased expression of Toll-like receptor-4 and MD-2 Spiegelberg HL, Horner AA, Takabayashi K, Raz E. Allergen- correlates with intestinal epithelial cell protection against immunostimulatory oligodeoxynucleotide conjugate: a novel dysregulated proinflammatory gene expression in response to allergoid for immunotherapy. Curr Opin Allergy Clin Immunol bacterial lipopolysaccharide. J Immunol 2001; 167: 1609-16.
90 Current Immunology Reviews, 2005, Vol. 1, No. 1
Bhattacharjee and Akira
Hausmann M, Kiessling S, Mestermann S, et al. Toll-like Furst DE, Lindsley H, Baethge B, et al. Dose-loading with receptors 2 and 4 are up-regulated during intestinal inflammation.
hydroxychloroquine improves the rate of response in early, active Gastroenterology 2002; 122: 1987-2000.
rheumatoid arthritis: a randomized, double-blind six-week trial Ogura Y, Bonen DK, Inohara N, et al. A frameshift mutation in with eighteen-week extension. Arthritis Rheum 1999; 42: 357-65.
NOD2 associated with susceptibility to Crohn's disease. Nature Leadbetter EA, Rifkin IR, Hohlbaum AM, Beaudette BC, Shlomchik MJ, Marshak-Rothstein A. Chromatin-IgG complexes Molad Y, Gorshtein A, Wysenbeek AJ, et al. Protective effect of activate B cells by dual engagement of IgM and Toll-like hydroxychloroquine in systemic lupus erythematosus. Prospective long-term study of an Israeli cohort. Lupus 2002; 11: 356-61.
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