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INNATE IMMUNITY 3
TLR3 Agonists
Poly(I:C) & Poly(I:C)-LMW
Polyinosine-polycytidylic acid (poly(I:C)) is a synthetic analog of double-stranded RNA (dsRNA), a molecular pattern associated with viral infection. Poly(I:C)
is recognized by TLR3 inducing the activation of NF-kB and the production of cytokines through distinct mechanisms that are MyD88-dependent or MyD88independent
1,2 . InvivoGen provides poly(I:C) with a high molecular weight (HMW) or a low molecular weight (LMW) that might activate the immune system
differently:
• Poly(I:C) (HMW) has an average size of 1.5-8 kb.
• Poly(I:C)-LMW has an average size of 0.2-1 kb.
Poly(A:U)
Polyadenylic–polyuridylic acid (poly(A:U)) is a synthetic double stranded RNA molecule that signals only through TLR3. Recognition of poly(A:U) by TLR3
induces the activation of dendritic cells and T lymphocytes. When combined with an antigen in mice, poly(A:U) has been shown to promote antigen-specific
Th1-immune responses and boost antibody production 3 . The potent adjuvant activity of poly(A:U) has been exploited in the treatment of breast cancers
that express TLR3 4 .
1. Yamamoto M. et al., 2003. Role of adaptor TRIF in the MyD88-independent toll-like receptor signaling pathway. Science, 301(5633):640-3. 2. Alexopoulou L. et al., 2001. Recognition of
double-stranded RNA and activation of NF-kappaB by Toll-like receptor 3. Nature, 413(6857):732-8. 3. Wang L. et al., 2002. Noncoding RNA danger motifs bridge innate and adaptive
immunity and are potent adjuvants for vaccination. J Clin Invest 110:1175–84. 4. Conforti R. et al., 2010. Opposing effects of toll-like receptor (TLR3) signaling in tumors can be therapeutically
uncoupled to optimize the anticancer efficacy of TLR3 ligands. Cancer Res. 70(2):490-500.
TLR4 Agonists
Bacterial lipopolysaccharide (LPS) is the major structural component of the outer wall of all Gram-negative bacteria and a potent activator of the immune
system. LPS is recognized by Toll-like receptor 4 (TLR4) which interacts with three different extracellular proteins: LPS binding protein (LBP), CD14 and,
myeloid differentiation protein 2 (MD-2), to induce a signaling cascade leading to the activation of NF-kB and the production of proinflammatory cytokines.
LPS consists of a polysaccharide region that is anchored in the outer bacterial membrane by a specific carbohydrate lipid moiety termed lipid A. Lipid A, also
known as endotoxin, is responsible for the immunostimulatory activity of LPS. The most active form of lipid A contains six fatty acyl groups and is found in
pathogenic bacteria such as Escherichia coli and Salmonella species 1 . Underacylated lipid A structures, containing four or five fatty acids, induce markedly less
host defense responses and can inhibit in a dose-dependent manner the strong endotoxic response triggered by hexa-acylated LPS 2 .
LPS-EB & LPS-EK standard (E. coli 0111:B4) - TLR4 (TLR2) Agonists
LPS-EB and LPS-EK are standard preparations of lipopolysaccharide. They are extracted by a phenol-water mixture. LPS-EB and LPS-EK contain other
bacterial components, such as lipopeptides, and therefore stimulate both TLR4 and TLR2.
LPS-EB, LPS-EK & LPS-SM Ultrapure (E. coli 0111:B4, E. coli K12 and S. minnesota)
Ultrapure LPS-EB (E. coli 0111:B4), LPS-EK (E. coli K12) and LPS-SM (S. minnesota Re type) are extracted by successive enzymatic hydrolysis steps and purified
by the phenol-TEA-DOC extraction protocol described by Hirschfeld M. et al. 3
LPS-RS (Rhodobacter sphaeroides) - TLR4 Antagonist
LPS from the photosynthetic bacterium Rhodobacter sphaeroides (LPS-RS) is a potent antagonist of LPS from pathogenic bacteria 1 . Complete competitive
inhibition of LPS activity is possible at a 100 fold excess of the antagonist. LPS-RS does not induce TLR4 signaling but is detected by the LAL assay, the
standard endotoxin detection assay.
MPLA & MPLAs (synthetic)
MPLA (monophosphoryl lipid A) is a derivative of lipid A from Salmonella minnesota R595 lipopolysaccharide (LPS or endotoxin). While LPS is a complex
heterogeneous molecule, its lipid A portion is relatively similar across a wide variety of pathogenic strains of bacteria 4 . MPLA, used extensively as a vaccine
adjuvant, has been shown to activate TLR4.
MPLAs is a synthetic monophosphoryl lipid A from E. coli with 6 fatty acyl groups. It is structurally very similar to natural MPLA except that natural MPLA
contains a mixture of 5, 6, and 7 acyl Lipid A. This E. coli synthetic MPLA activates TLR4 but does not activate TLR2 even at high concentrations reflecting its
high purity.
1. Coats SR. et al., 2005. MD-2 mediates the ability of tetra-acylated and penta-acylated lipopolysaccharides to antagonize Escherichia coli lipopolysaccharide at the TLR4 signaling complex.
J Immunol.;175(7):4490-8. 2. Teghanemt A. et al., 2005. Molecular basis of reduced potency of underacylated endotoxins. J Immunol. 175(7):4669-76. 3. Hirschfeld M. et al., 2000. Cutting edge:
repurification of lipopolysaccharide eliminates signaling through both human and murine toll-like receptor 2. J Immunol. ;165(2):618-22. 4. Martin M. et al., 2003. Role of innate immune
factors in the adjuvant activity of monophosphoryl lipid A. Infect Immun. 71(5):2498-507.
TLR5 Agonists
FLA-BS, FLA-ST, FLA-ST Ultrapure & RecFLA-ST (B. subtilis and S. typhimurium)
Flagellin is the major component of the bacterial flagellar filament, which confers motility on a wide range of bacterial species. Flagellin is recognized by TLR5 1
and induces the activation of NF-kB and the production of cytokines and nitric oxide depending on the nature of the TLR5 signaling complex 2 .
• FLA-BS and FLA-ST are flagellins isolated from the Gram-positive bacteria B. subtilis and from the Gram-negative bacteria S. typhimurium, respectively. They
are purified by acid hydrolysis, heating and ultrafiltration according to Ibrahim GF. et al. 3 . The purity of FLA-ST is estimated at 10%.
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