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Small-molecule activators of insulin-degrading enzyme discovered through high-throughput compound screening.

Cabrol C, Huzarska MA, Dinolfo C, Rodriguez MC, Reinstatler L, Ni J, Yeh LA, Cuny GD, Stein RL, Selkoe DJ, Leissring MA - PLoS ONE (2009)

Bottom Line: Both compounds were found to interfere with the crosslinking of a photoaffinity ATP analogue to IDE, suggesting that they interact with a bona fide ATP-binding domain within IDE.These novel activators help to establish the putative ATP-binding domain as a key modulator of IDE proteolytic activity and offer new insights into the modulatory action of ATP.Several larger lessons abstracted from this screen will help inform the design of future screening campaigns and facilitate the eventual development of IDE activators with therapeutic utility.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, Mayo Clinic, Jacksonville, Florida, United States of America.

ABSTRACT

Background: Hypocatabolism of the amyloid beta-protein (Abeta) by insulin-degrading enzyme (IDE) is implicated in the pathogenesis of Alzheimer disease (AD), making pharmacological activation of IDE an attractive therapeutic strategy. However, it has not been established whether the proteolytic activity of IDE can be enhanced by drug-like compounds.

Methodology/principal findings: Based on the finding that ATP and other nucleotide polyphosphates modulate IDE activity at physiological concentrations, we conducted parallel high-throughput screening campaigns in the absence or presence of ATP and identified two compounds--designated Ia1 and Ia2--that significantly stimulate IDE proteolytic activity. Both compounds were found to interfere with the crosslinking of a photoaffinity ATP analogue to IDE, suggesting that they interact with a bona fide ATP-binding domain within IDE. Unexpectedly, we observed highly synergistic activation effects when the activity of Ia1 or Ia2 was tested in the presence of ATP, a finding that has implications for the mechanisms underlying ATP-mediated activation of IDE. Notably, Ia1 and Ia2 activated the degradation of Abeta by approximately 700% and approximately 400%, respectively, albeit only when Abeta was presented in a mixture also containing shorter substrates.

Conclusions/significance: This study describes the first examples of synthetic small-molecule activators of IDE, showing that pharmacological activation of this important protease with drug-like compounds is achievable. These novel activators help to establish the putative ATP-binding domain as a key modulator of IDE proteolytic activity and offer new insights into the modulatory action of ATP. Several larger lessons abstracted from this screen will help inform the design of future screening campaigns and facilitate the eventual development of IDE activators with therapeutic utility.

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Related in: MedlinePlus

Small-molecule activators of IDE discovered through compound screening.A,B, Structures of Ia1 (A) and Ia2 (B) and dose-dependent effects on the IDE-mediated hydrolysis of SP1 and FRET1. Data are mean±SEM for 4 to 5 independent experiments normalized to control wells containing no ATP. Data for all doses showing ≥30% activation are statistically different from DMSO controls (P<0.05).
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pone-0005274-g002: Small-molecule activators of IDE discovered through compound screening.A,B, Structures of Ia1 (A) and Ia2 (B) and dose-dependent effects on the IDE-mediated hydrolysis of SP1 and FRET1. Data are mean±SEM for 4 to 5 independent experiments normalized to control wells containing no ATP. Data for all doses showing ≥30% activation are statistically different from DMSO controls (P<0.05).

Mentions: Two compounds that consistently activated IDE—dubbed Ia1 and Ia2 (IDE activators 1 and 2; LDN-1487 and LDN-1844, respectively)—emerged from the screening campaign (Fig. 2A,B). Ia1 activated the hydrolysis of SP1 and FRET1 by up to ∼200% and ∼500%, respectively, in a dose-dependent manner, with maximal activation occurring at 200 µM (Fig. 2A). Ia2 also activated both substrates, albeit to a lesser extent: maximal activation by Ia2 was determined to be ∼110% and ∼60% at concentrations of 50 µM and 6.25 µM, for SP1 and FRET1, respectively.


Small-molecule activators of insulin-degrading enzyme discovered through high-throughput compound screening.

Cabrol C, Huzarska MA, Dinolfo C, Rodriguez MC, Reinstatler L, Ni J, Yeh LA, Cuny GD, Stein RL, Selkoe DJ, Leissring MA - PLoS ONE (2009)

Small-molecule activators of IDE discovered through compound screening.A,B, Structures of Ia1 (A) and Ia2 (B) and dose-dependent effects on the IDE-mediated hydrolysis of SP1 and FRET1. Data are mean±SEM for 4 to 5 independent experiments normalized to control wells containing no ATP. Data for all doses showing ≥30% activation are statistically different from DMSO controls (P<0.05).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2668070&req=5

pone-0005274-g002: Small-molecule activators of IDE discovered through compound screening.A,B, Structures of Ia1 (A) and Ia2 (B) and dose-dependent effects on the IDE-mediated hydrolysis of SP1 and FRET1. Data are mean±SEM for 4 to 5 independent experiments normalized to control wells containing no ATP. Data for all doses showing ≥30% activation are statistically different from DMSO controls (P<0.05).
Mentions: Two compounds that consistently activated IDE—dubbed Ia1 and Ia2 (IDE activators 1 and 2; LDN-1487 and LDN-1844, respectively)—emerged from the screening campaign (Fig. 2A,B). Ia1 activated the hydrolysis of SP1 and FRET1 by up to ∼200% and ∼500%, respectively, in a dose-dependent manner, with maximal activation occurring at 200 µM (Fig. 2A). Ia2 also activated both substrates, albeit to a lesser extent: maximal activation by Ia2 was determined to be ∼110% and ∼60% at concentrations of 50 µM and 6.25 µM, for SP1 and FRET1, respectively.

Bottom Line: Both compounds were found to interfere with the crosslinking of a photoaffinity ATP analogue to IDE, suggesting that they interact with a bona fide ATP-binding domain within IDE.These novel activators help to establish the putative ATP-binding domain as a key modulator of IDE proteolytic activity and offer new insights into the modulatory action of ATP.Several larger lessons abstracted from this screen will help inform the design of future screening campaigns and facilitate the eventual development of IDE activators with therapeutic utility.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, Mayo Clinic, Jacksonville, Florida, United States of America.

ABSTRACT

Background: Hypocatabolism of the amyloid beta-protein (Abeta) by insulin-degrading enzyme (IDE) is implicated in the pathogenesis of Alzheimer disease (AD), making pharmacological activation of IDE an attractive therapeutic strategy. However, it has not been established whether the proteolytic activity of IDE can be enhanced by drug-like compounds.

Methodology/principal findings: Based on the finding that ATP and other nucleotide polyphosphates modulate IDE activity at physiological concentrations, we conducted parallel high-throughput screening campaigns in the absence or presence of ATP and identified two compounds--designated Ia1 and Ia2--that significantly stimulate IDE proteolytic activity. Both compounds were found to interfere with the crosslinking of a photoaffinity ATP analogue to IDE, suggesting that they interact with a bona fide ATP-binding domain within IDE. Unexpectedly, we observed highly synergistic activation effects when the activity of Ia1 or Ia2 was tested in the presence of ATP, a finding that has implications for the mechanisms underlying ATP-mediated activation of IDE. Notably, Ia1 and Ia2 activated the degradation of Abeta by approximately 700% and approximately 400%, respectively, albeit only when Abeta was presented in a mixture also containing shorter substrates.

Conclusions/significance: This study describes the first examples of synthetic small-molecule activators of IDE, showing that pharmacological activation of this important protease with drug-like compounds is achievable. These novel activators help to establish the putative ATP-binding domain as a key modulator of IDE proteolytic activity and offer new insights into the modulatory action of ATP. Several larger lessons abstracted from this screen will help inform the design of future screening campaigns and facilitate the eventual development of IDE activators with therapeutic utility.

Show MeSH
Related in: MedlinePlus