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Receptor residence time trumps drug-likeness and oral bioavailability in determining efficacy of complement C5a antagonists.

Seow V, Lim J, Cotterell AJ, Yau MK, Xu W, Lohman RJ, Kok WM, Stoermer MJ, Sweet MJ, Reid RC, Suen JY, Fairlie DP - Sci Rep (2016)

Bottom Line: The unusually long residence time of 3D53 on its receptor was mechanistically probed by molecular dynamics simulations, which revealed long-lasting interactions that trap the antagonist within the receptor.Despite negligible oral bioavailability, 3D53 was much more orally efficacious than W54011 or JJ47 in preventing repeated agonist insults to induce rat paw oedema over 24 h.Thus, residence time on a receptor can trump drug-likeness in determining efficacy, even oral efficacy, of pharmacological agents.

View Article: PubMed Central - PubMed

Affiliation: Division of Chemistry and Structural Biology, Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072, Australia.

ABSTRACT
Drug discovery and translation are normally based on optimizing efficacy by increasing receptor affinity, functional potency, drug-likeness (rule-of-five compliance) and oral bioavailability. Here we demonstrate that residence time of a compound on its receptor has an overriding influence on efficacy, exemplified for antagonists of inflammatory protein complement C5a that activates immune cells and promotes disease. Three equipotent antagonists (3D53, W54011, JJ47) of inflammatory responses to C5a (3 nM) were compared for drug-likeness, receptor affinity and antagonist potency in human macrophages, and anti-inflammatory efficacy in rats. Only the least drug-like antagonist (3D53) maintained potency in cells against higher C5a concentrations and had a much longer duration of action (t1/2 ~ 20 h) than W54011 or JJ47 (t1/2 ~ 1 -3 h) in inhibiting macrophage responses. The unusually long residence time of 3D53 on its receptor was mechanistically probed by molecular dynamics simulations, which revealed long-lasting interactions that trap the antagonist within the receptor. Despite negligible oral bioavailability, 3D53 was much more orally efficacious than W54011 or JJ47 in preventing repeated agonist insults to induce rat paw oedema over 24 h. Thus, residence time on a receptor can trump drug-likeness in determining efficacy, even oral efficacy, of pharmacological agents.

No MeSH data available.


Related in: MedlinePlus

3D53 is more effective than W54011 and JJ47 in inhibiting C5a-induced migration or calcium release for human HMDM due to longer residence time on C5aR.(A) Migration of HMDM through 5 μm Transwell inserts with or without 3 nM C5a for 16 h. For antagonism of C5aR, cells were treated with 3D53 (0.1 or 1 μM), W54011 (0.1 or 1 μM) or JJ47 (0.1 or 1 μM) for 1 h. Unbound antagonists were washed away prior to stimulation with 3 nM C5a. Chemotactic index is expressed as fold change against control. (B) C5a induced Ca2+ release from HMDM, pre-treated with 1 μM C5aR antagonist (●) 3D53, (▲) W54011 and (○) JJ47 for 1 h. Excess unbound antagonists were removed by washing. Antagonist residence time on C5aR was determined by subjecting treated cells to 3 nM C5a during the stated duration post antagonist-treatment up to 4 h and (C) up to 60 h for 3D53 in a calcium release assay. Calculated half-life for 3D53, W54011 and JJ47 are 18.2 h, 1.2 h and 0.6 h respectively. Error bars are means ± SEM of three independent experiments (n = 3). ***p < 0.001 by student t-test.
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f4: 3D53 is more effective than W54011 and JJ47 in inhibiting C5a-induced migration or calcium release for human HMDM due to longer residence time on C5aR.(A) Migration of HMDM through 5 μm Transwell inserts with or without 3 nM C5a for 16 h. For antagonism of C5aR, cells were treated with 3D53 (0.1 or 1 μM), W54011 (0.1 or 1 μM) or JJ47 (0.1 or 1 μM) for 1 h. Unbound antagonists were washed away prior to stimulation with 3 nM C5a. Chemotactic index is expressed as fold change against control. (B) C5a induced Ca2+ release from HMDM, pre-treated with 1 μM C5aR antagonist (●) 3D53, (▲) W54011 and (○) JJ47 for 1 h. Excess unbound antagonists were removed by washing. Antagonist residence time on C5aR was determined by subjecting treated cells to 3 nM C5a during the stated duration post antagonist-treatment up to 4 h and (C) up to 60 h for 3D53 in a calcium release assay. Calculated half-life for 3D53, W54011 and JJ47 are 18.2 h, 1.2 h and 0.6 h respectively. Error bars are means ± SEM of three independent experiments (n = 3). ***p < 0.001 by student t-test.

Mentions: Insurmountable antagonists with a long residence time (slow off-rate) from the receptor-binding site may not reach a true equilibrium between the agonist and antagonist on the receptor in the short timeframe of the calcium release assay (i.e. <5 min). Since C5a is one of the most potent endogenous agents known to induce chemotaxis of immune cells, a more stringent and longer duration test of surmountability is a chemotaxis migration assay, which was performed to investigate the C5aR antagonist mechanism. The antagonist-treated HMDM were exposed to C5a for 16 h continuously. Antagonists with a short residence time will be ineffective in this assay, because once the antagonist has dissociated from C5aR, C5a immediately binds and induces migration. While 3D53 was able to block C5a-induced migration at a concentration of 100 nM, W54011 and JJ47 were not effective antagonists even at 1 μM concentrations (Fig. 4A) over this 16 h timeframe. Since W54011 and JJ47 failed to block chemotaxis here, it seems unlikely that these antagonists would be functional in vivo in blocking chemotaxis.


Receptor residence time trumps drug-likeness and oral bioavailability in determining efficacy of complement C5a antagonists.

Seow V, Lim J, Cotterell AJ, Yau MK, Xu W, Lohman RJ, Kok WM, Stoermer MJ, Sweet MJ, Reid RC, Suen JY, Fairlie DP - Sci Rep (2016)

3D53 is more effective than W54011 and JJ47 in inhibiting C5a-induced migration or calcium release for human HMDM due to longer residence time on C5aR.(A) Migration of HMDM through 5 μm Transwell inserts with or without 3 nM C5a for 16 h. For antagonism of C5aR, cells were treated with 3D53 (0.1 or 1 μM), W54011 (0.1 or 1 μM) or JJ47 (0.1 or 1 μM) for 1 h. Unbound antagonists were washed away prior to stimulation with 3 nM C5a. Chemotactic index is expressed as fold change against control. (B) C5a induced Ca2+ release from HMDM, pre-treated with 1 μM C5aR antagonist (●) 3D53, (▲) W54011 and (○) JJ47 for 1 h. Excess unbound antagonists were removed by washing. Antagonist residence time on C5aR was determined by subjecting treated cells to 3 nM C5a during the stated duration post antagonist-treatment up to 4 h and (C) up to 60 h for 3D53 in a calcium release assay. Calculated half-life for 3D53, W54011 and JJ47 are 18.2 h, 1.2 h and 0.6 h respectively. Error bars are means ± SEM of three independent experiments (n = 3). ***p < 0.001 by student t-test.
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f4: 3D53 is more effective than W54011 and JJ47 in inhibiting C5a-induced migration or calcium release for human HMDM due to longer residence time on C5aR.(A) Migration of HMDM through 5 μm Transwell inserts with or without 3 nM C5a for 16 h. For antagonism of C5aR, cells were treated with 3D53 (0.1 or 1 μM), W54011 (0.1 or 1 μM) or JJ47 (0.1 or 1 μM) for 1 h. Unbound antagonists were washed away prior to stimulation with 3 nM C5a. Chemotactic index is expressed as fold change against control. (B) C5a induced Ca2+ release from HMDM, pre-treated with 1 μM C5aR antagonist (●) 3D53, (▲) W54011 and (○) JJ47 for 1 h. Excess unbound antagonists were removed by washing. Antagonist residence time on C5aR was determined by subjecting treated cells to 3 nM C5a during the stated duration post antagonist-treatment up to 4 h and (C) up to 60 h for 3D53 in a calcium release assay. Calculated half-life for 3D53, W54011 and JJ47 are 18.2 h, 1.2 h and 0.6 h respectively. Error bars are means ± SEM of three independent experiments (n = 3). ***p < 0.001 by student t-test.
Mentions: Insurmountable antagonists with a long residence time (slow off-rate) from the receptor-binding site may not reach a true equilibrium between the agonist and antagonist on the receptor in the short timeframe of the calcium release assay (i.e. <5 min). Since C5a is one of the most potent endogenous agents known to induce chemotaxis of immune cells, a more stringent and longer duration test of surmountability is a chemotaxis migration assay, which was performed to investigate the C5aR antagonist mechanism. The antagonist-treated HMDM were exposed to C5a for 16 h continuously. Antagonists with a short residence time will be ineffective in this assay, because once the antagonist has dissociated from C5aR, C5a immediately binds and induces migration. While 3D53 was able to block C5a-induced migration at a concentration of 100 nM, W54011 and JJ47 were not effective antagonists even at 1 μM concentrations (Fig. 4A) over this 16 h timeframe. Since W54011 and JJ47 failed to block chemotaxis here, it seems unlikely that these antagonists would be functional in vivo in blocking chemotaxis.

Bottom Line: The unusually long residence time of 3D53 on its receptor was mechanistically probed by molecular dynamics simulations, which revealed long-lasting interactions that trap the antagonist within the receptor.Despite negligible oral bioavailability, 3D53 was much more orally efficacious than W54011 or JJ47 in preventing repeated agonist insults to induce rat paw oedema over 24 h.Thus, residence time on a receptor can trump drug-likeness in determining efficacy, even oral efficacy, of pharmacological agents.

View Article: PubMed Central - PubMed

Affiliation: Division of Chemistry and Structural Biology, Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072, Australia.

ABSTRACT
Drug discovery and translation are normally based on optimizing efficacy by increasing receptor affinity, functional potency, drug-likeness (rule-of-five compliance) and oral bioavailability. Here we demonstrate that residence time of a compound on its receptor has an overriding influence on efficacy, exemplified for antagonists of inflammatory protein complement C5a that activates immune cells and promotes disease. Three equipotent antagonists (3D53, W54011, JJ47) of inflammatory responses to C5a (3 nM) were compared for drug-likeness, receptor affinity and antagonist potency in human macrophages, and anti-inflammatory efficacy in rats. Only the least drug-like antagonist (3D53) maintained potency in cells against higher C5a concentrations and had a much longer duration of action (t1/2 ~ 20 h) than W54011 or JJ47 (t1/2 ~ 1 -3 h) in inhibiting macrophage responses. The unusually long residence time of 3D53 on its receptor was mechanistically probed by molecular dynamics simulations, which revealed long-lasting interactions that trap the antagonist within the receptor. Despite negligible oral bioavailability, 3D53 was much more orally efficacious than W54011 or JJ47 in preventing repeated agonist insults to induce rat paw oedema over 24 h. Thus, residence time on a receptor can trump drug-likeness in determining efficacy, even oral efficacy, of pharmacological agents.

No MeSH data available.


Related in: MedlinePlus