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Glycosaminoglycan analogs as a novel anti-inflammatory strategy.

Severin IC, Soares A, Hantson J, Teixeira M, Sachs D, Valognes D, Scheer A, Schwarz MK, Wells TN, Proudfoot AE, Shaw J - Front Immunol (2012)

Bottom Line: In vitro, these molecules prevented chemokine-GAG binding and chemokine receptor activation without disrupting coagulation.However, in vivo, these compounds caused variable results in a murine peritoneal recruitment assay, with a general increase of cell recruitment.In more disease specific models, such as antigen-induced arthritis and delayed-type hypersensitivity, an overall decrease in inflammation was noted, suggesting that the primary anti-inflammatory effect may also involve factors beyond the chemokine system.

View Article: PubMed Central - PubMed

Affiliation: Merck Serono Geneva Research Centre Geneva, Switzerland.

ABSTRACT
Heparin, a glycosaminoglycan (GAG), has both anti-inflammatory and anti-coagulant properties. The clinical use of heparin against inflammation, however, has been limited by concerns about increased bleeding. While the anti-coagulant activity of heparin is well understood, its anti-inflammatory properties are less so. Heparin is known to bind to certain cytokines, including chemokines, small proteins which mediate inflammation through their control of leukocyte migration and activation. Molecules which can interrupt the chemokine-GAG interaction without inhibiting coagulation could therefore, represent a new class of anti-inflammatory agents. In the present study, two approaches were undertaken, both focusing on the heparin-chemokine relationship. In the first, a structure based strategy was used: after an initial screening of potential small molecule binders using protein NMR on a target chemokine, binding molecules were optimized through structure-based design. In the second approach, commercially available short oligosaccharides were polysulfated. In vitro, these molecules prevented chemokine-GAG binding and chemokine receptor activation without disrupting coagulation. However, in vivo, these compounds caused variable results in a murine peritoneal recruitment assay, with a general increase of cell recruitment. In more disease specific models, such as antigen-induced arthritis and delayed-type hypersensitivity, an overall decrease in inflammation was noted, suggesting that the primary anti-inflammatory effect may also involve factors beyond the chemokine system.

No MeSH data available.


Related in: MedlinePlus

Inhibition of CCL5 binding by GAG analogs. Inhibition of CCL5 binding to heparin was measured by the ability to compete for 125I-CCL5 binding to heparin beads by Molecule 1 (A) and persulfated oligosaccharides (C). Inhibition of CCL5 binding to its receptors was determined by competition equilibrium binding assays using membranes from transfectants expressing CCR1 (B) and (D) or CCR5 (E) by persulfated oligosaccharides. Legend: • Molecule 1; ■ Molecule 3; □ Molecule 1 on CCR5; ♦ MHxS; ▼ StachS; ▲ NisS; ○, 3 kDa (H3400) heparin.
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Figure 3: Inhibition of CCL5 binding by GAG analogs. Inhibition of CCL5 binding to heparin was measured by the ability to compete for 125I-CCL5 binding to heparin beads by Molecule 1 (A) and persulfated oligosaccharides (C). Inhibition of CCL5 binding to its receptors was determined by competition equilibrium binding assays using membranes from transfectants expressing CCR1 (B) and (D) or CCR5 (E) by persulfated oligosaccharides. Legend: • Molecule 1; ■ Molecule 3; □ Molecule 1 on CCR5; ♦ MHxS; ▼ StachS; ▲ NisS; ○, 3 kDa (H3400) heparin.

Mentions: A study of the relative affinity of Molecule 1 toward CCL5 and heparin molecules was first evaluated by competition experiments in which CCL5, immobilized on heparin beads, was competed off the beads by increasing concentrations of Molecule 1. In this experiment, an IC50 of 0.32 mM for Molecule 1 was observed (Figure 3A). Despite this low IC50, Molecule 1 is the first small molecule identified capable of disrupting the interaction between heparin and CCL5. Since the principal GAG binding motif of CCL5, the BBXB motif on the 40s loop, plays a role in binding to CCR1, we determined the capacity of Molecule 1 to inhibit the binding of the chemokine CCL5 to its receptors CCR1 and CCR5 using equilibrium competition binding with a SPA-based assay. Molecule 1 was in fact more potent in this assay, as it competed 125CCL5 for binding to CHO/CCR1 transfectants with an IC50 value of 6.7 μM, but had no effect on CCL5 binding to CCR5, consistent with the fact that the GAG binding motifs of CCL5 are distinct from those for CCR5 (Figure 3B). In order to evaluate if Molecule 1 could be a general CCR1 antagonist, the same experiment was performed with CCL3 as the ligand; however, no competition of CCL3 to its receptors CCR1 and CCR5 was observed (data not shown). We conclude that Molecule 1 is selective for CCL5 binding to CCR1. However, contrary to our predictions, Molecule 3 demonstrated no increased potency as it demonstrated an IC50 value of 2.5 μM for inhibition of binding to CCR1 (Table 1, Figure 3B).


Glycosaminoglycan analogs as a novel anti-inflammatory strategy.

Severin IC, Soares A, Hantson J, Teixeira M, Sachs D, Valognes D, Scheer A, Schwarz MK, Wells TN, Proudfoot AE, Shaw J - Front Immunol (2012)

Inhibition of CCL5 binding by GAG analogs. Inhibition of CCL5 binding to heparin was measured by the ability to compete for 125I-CCL5 binding to heparin beads by Molecule 1 (A) and persulfated oligosaccharides (C). Inhibition of CCL5 binding to its receptors was determined by competition equilibrium binding assays using membranes from transfectants expressing CCR1 (B) and (D) or CCR5 (E) by persulfated oligosaccharides. Legend: • Molecule 1; ■ Molecule 3; □ Molecule 1 on CCR5; ♦ MHxS; ▼ StachS; ▲ NisS; ○, 3 kDa (H3400) heparin.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3472544&req=5

Figure 3: Inhibition of CCL5 binding by GAG analogs. Inhibition of CCL5 binding to heparin was measured by the ability to compete for 125I-CCL5 binding to heparin beads by Molecule 1 (A) and persulfated oligosaccharides (C). Inhibition of CCL5 binding to its receptors was determined by competition equilibrium binding assays using membranes from transfectants expressing CCR1 (B) and (D) or CCR5 (E) by persulfated oligosaccharides. Legend: • Molecule 1; ■ Molecule 3; □ Molecule 1 on CCR5; ♦ MHxS; ▼ StachS; ▲ NisS; ○, 3 kDa (H3400) heparin.
Mentions: A study of the relative affinity of Molecule 1 toward CCL5 and heparin molecules was first evaluated by competition experiments in which CCL5, immobilized on heparin beads, was competed off the beads by increasing concentrations of Molecule 1. In this experiment, an IC50 of 0.32 mM for Molecule 1 was observed (Figure 3A). Despite this low IC50, Molecule 1 is the first small molecule identified capable of disrupting the interaction between heparin and CCL5. Since the principal GAG binding motif of CCL5, the BBXB motif on the 40s loop, plays a role in binding to CCR1, we determined the capacity of Molecule 1 to inhibit the binding of the chemokine CCL5 to its receptors CCR1 and CCR5 using equilibrium competition binding with a SPA-based assay. Molecule 1 was in fact more potent in this assay, as it competed 125CCL5 for binding to CHO/CCR1 transfectants with an IC50 value of 6.7 μM, but had no effect on CCL5 binding to CCR5, consistent with the fact that the GAG binding motifs of CCL5 are distinct from those for CCR5 (Figure 3B). In order to evaluate if Molecule 1 could be a general CCR1 antagonist, the same experiment was performed with CCL3 as the ligand; however, no competition of CCL3 to its receptors CCR1 and CCR5 was observed (data not shown). We conclude that Molecule 1 is selective for CCL5 binding to CCR1. However, contrary to our predictions, Molecule 3 demonstrated no increased potency as it demonstrated an IC50 value of 2.5 μM for inhibition of binding to CCR1 (Table 1, Figure 3B).

Bottom Line: In vitro, these molecules prevented chemokine-GAG binding and chemokine receptor activation without disrupting coagulation.However, in vivo, these compounds caused variable results in a murine peritoneal recruitment assay, with a general increase of cell recruitment.In more disease specific models, such as antigen-induced arthritis and delayed-type hypersensitivity, an overall decrease in inflammation was noted, suggesting that the primary anti-inflammatory effect may also involve factors beyond the chemokine system.

View Article: PubMed Central - PubMed

Affiliation: Merck Serono Geneva Research Centre Geneva, Switzerland.

ABSTRACT
Heparin, a glycosaminoglycan (GAG), has both anti-inflammatory and anti-coagulant properties. The clinical use of heparin against inflammation, however, has been limited by concerns about increased bleeding. While the anti-coagulant activity of heparin is well understood, its anti-inflammatory properties are less so. Heparin is known to bind to certain cytokines, including chemokines, small proteins which mediate inflammation through their control of leukocyte migration and activation. Molecules which can interrupt the chemokine-GAG interaction without inhibiting coagulation could therefore, represent a new class of anti-inflammatory agents. In the present study, two approaches were undertaken, both focusing on the heparin-chemokine relationship. In the first, a structure based strategy was used: after an initial screening of potential small molecule binders using protein NMR on a target chemokine, binding molecules were optimized through structure-based design. In the second approach, commercially available short oligosaccharides were polysulfated. In vitro, these molecules prevented chemokine-GAG binding and chemokine receptor activation without disrupting coagulation. However, in vivo, these compounds caused variable results in a murine peritoneal recruitment assay, with a general increase of cell recruitment. In more disease specific models, such as antigen-induced arthritis and delayed-type hypersensitivity, an overall decrease in inflammation was noted, suggesting that the primary anti-inflammatory effect may also involve factors beyond the chemokine system.

No MeSH data available.


Related in: MedlinePlus