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Induction of autophagy by a novel small molecule improves aβ pathology and ameliorates cognitive deficits.

Chu C, Zhang X, Ma W, Li L, Wang W, Shang L, Fu P - PLoS ONE (2013)

Bottom Line: GTM-1 modulates autophagy in an Akt-independent and mTOR-independent manner.In addition, we demonstrated that GTM-1 enhances autophagy clearance and reverses the downregulation of autophagy flux by thapsigargin and asparagine.Furthermore, administration of GTM-1 attenuated Aβ pathology and ameliorated cognitive deficits in AD mice.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, The First People's Hospital of Yangzhou, Jiang Su, PR China.

ABSTRACT
Growing evidence has demonstrated a neuroprotective role of autophagy in Alzheimer's disease (AD). Thus, autophagy has been regarded as a potential therapeutic target, attracting increasing interest in pharmaceutical autophagy modulation by small molecules. We designed a two-cycle screening strategy on the basis of imaging high-throughout screening (HTS) and cellular toxicity assay, and have identified a novel autophagy inducer known as GTM-1. We further showed that GTM-1 exhibits dual activities, such as autophagy induction and antagonism against Aβ-oligomer toxicity. GTM-1 modulates autophagy in an Akt-independent and mTOR-independent manner. In addition, we demonstrated that GTM-1 enhances autophagy clearance and reverses the downregulation of autophagy flux by thapsigargin and asparagine. Furthermore, administration of GTM-1 attenuated Aβ pathology and ameliorated cognitive deficits in AD mice.

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Isolation of a small-molecule inducer of autophagy in neurons.A, The structure of GTM-1. B–D, GTM-1 increases the spot intensity of LC3-GFP+ puncta. SH-SY5Y/LC3-GFP cells were treated with 20 µM GTM-1 for the indicated times (B) or with the indicated concentrations (C) for 24 hrs. The spot intensity of LC3-GFP+ puncta in SH-SY5Y/LC3-GFP cells treated with control vehicle at first time point (0 hr) was used as the negative control (NC). The image data were expressed as % of NC. For each treatment condition, 1000 cells were analyzed. The cells were imaged using a fluorescence microscope (D). E, SH-SY5Y cells were incubated with the indicated compounds for 24 hrs. The cell viability was assayed using the MTT assay. Aβ, Aβ42 (30 µM). Cells treated with vehicle (0.1% DMSO) were used as a negative control (NC).
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pone-0065367-g001: Isolation of a small-molecule inducer of autophagy in neurons.A, The structure of GTM-1. B–D, GTM-1 increases the spot intensity of LC3-GFP+ puncta. SH-SY5Y/LC3-GFP cells were treated with 20 µM GTM-1 for the indicated times (B) or with the indicated concentrations (C) for 24 hrs. The spot intensity of LC3-GFP+ puncta in SH-SY5Y/LC3-GFP cells treated with control vehicle at first time point (0 hr) was used as the negative control (NC). The image data were expressed as % of NC. For each treatment condition, 1000 cells were analyzed. The cells were imaged using a fluorescence microscope (D). E, SH-SY5Y cells were incubated with the indicated compounds for 24 hrs. The cell viability was assayed using the MTT assay. Aβ, Aβ42 (30 µM). Cells treated with vehicle (0.1% DMSO) were used as a negative control (NC).

Mentions: A previous report conducted by Hung et al. adopted extracellular Aβ to mimic the brain morphological damages caused by fibrillogenic Aβ at the cellular level. It has been demonstrated that extracellular Aβ can be easily internalized, causing neuronal apoptosis after simple incubation [29]. On the basis of this finding, we performed an Aβ-induced cytotoxicity test. From this study, 15 hits were further examined using the cellular Aβ-induced cytotoxicity test [29] in the second screening (studies examining the roles of these other hits are still ongoing). Using this assay to examine the extent of neuroprotection offered by these compounds, we identified a quinazoline derivative, GTM-1 (Fig. 1A), which exhibited dual activities in autophagy induction and antagonism against Aβ-oligomer toxicity (Fig. 1E). The NMR spectrum of GTM-1 is shown in figure 1 (Fig. 1). A quantitative analysis of the LC3-GFP puncta induced by GTM-1 (Fig. 1C) using HCS demonstrated that the autophagy induction of GTM-1 in SH-SY5Y/LC3-GFP cells was as rapid as 4 hrs (Fig. 1B) and occurred in a dose-dependent manner with an EC50 of 7.9 µM (Fig. 1C).


Induction of autophagy by a novel small molecule improves aβ pathology and ameliorates cognitive deficits.

Chu C, Zhang X, Ma W, Li L, Wang W, Shang L, Fu P - PLoS ONE (2013)

Isolation of a small-molecule inducer of autophagy in neurons.A, The structure of GTM-1. B–D, GTM-1 increases the spot intensity of LC3-GFP+ puncta. SH-SY5Y/LC3-GFP cells were treated with 20 µM GTM-1 for the indicated times (B) or with the indicated concentrations (C) for 24 hrs. The spot intensity of LC3-GFP+ puncta in SH-SY5Y/LC3-GFP cells treated with control vehicle at first time point (0 hr) was used as the negative control (NC). The image data were expressed as % of NC. For each treatment condition, 1000 cells were analyzed. The cells were imaged using a fluorescence microscope (D). E, SH-SY5Y cells were incubated with the indicated compounds for 24 hrs. The cell viability was assayed using the MTT assay. Aβ, Aβ42 (30 µM). Cells treated with vehicle (0.1% DMSO) were used as a negative control (NC).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3672196&req=5

pone-0065367-g001: Isolation of a small-molecule inducer of autophagy in neurons.A, The structure of GTM-1. B–D, GTM-1 increases the spot intensity of LC3-GFP+ puncta. SH-SY5Y/LC3-GFP cells were treated with 20 µM GTM-1 for the indicated times (B) or with the indicated concentrations (C) for 24 hrs. The spot intensity of LC3-GFP+ puncta in SH-SY5Y/LC3-GFP cells treated with control vehicle at first time point (0 hr) was used as the negative control (NC). The image data were expressed as % of NC. For each treatment condition, 1000 cells were analyzed. The cells were imaged using a fluorescence microscope (D). E, SH-SY5Y cells were incubated with the indicated compounds for 24 hrs. The cell viability was assayed using the MTT assay. Aβ, Aβ42 (30 µM). Cells treated with vehicle (0.1% DMSO) were used as a negative control (NC).
Mentions: A previous report conducted by Hung et al. adopted extracellular Aβ to mimic the brain morphological damages caused by fibrillogenic Aβ at the cellular level. It has been demonstrated that extracellular Aβ can be easily internalized, causing neuronal apoptosis after simple incubation [29]. On the basis of this finding, we performed an Aβ-induced cytotoxicity test. From this study, 15 hits were further examined using the cellular Aβ-induced cytotoxicity test [29] in the second screening (studies examining the roles of these other hits are still ongoing). Using this assay to examine the extent of neuroprotection offered by these compounds, we identified a quinazoline derivative, GTM-1 (Fig. 1A), which exhibited dual activities in autophagy induction and antagonism against Aβ-oligomer toxicity (Fig. 1E). The NMR spectrum of GTM-1 is shown in figure 1 (Fig. 1). A quantitative analysis of the LC3-GFP puncta induced by GTM-1 (Fig. 1C) using HCS demonstrated that the autophagy induction of GTM-1 in SH-SY5Y/LC3-GFP cells was as rapid as 4 hrs (Fig. 1B) and occurred in a dose-dependent manner with an EC50 of 7.9 µM (Fig. 1C).

Bottom Line: GTM-1 modulates autophagy in an Akt-independent and mTOR-independent manner.In addition, we demonstrated that GTM-1 enhances autophagy clearance and reverses the downregulation of autophagy flux by thapsigargin and asparagine.Furthermore, administration of GTM-1 attenuated Aβ pathology and ameliorated cognitive deficits in AD mice.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, The First People's Hospital of Yangzhou, Jiang Su, PR China.

ABSTRACT
Growing evidence has demonstrated a neuroprotective role of autophagy in Alzheimer's disease (AD). Thus, autophagy has been regarded as a potential therapeutic target, attracting increasing interest in pharmaceutical autophagy modulation by small molecules. We designed a two-cycle screening strategy on the basis of imaging high-throughout screening (HTS) and cellular toxicity assay, and have identified a novel autophagy inducer known as GTM-1. We further showed that GTM-1 exhibits dual activities, such as autophagy induction and antagonism against Aβ-oligomer toxicity. GTM-1 modulates autophagy in an Akt-independent and mTOR-independent manner. In addition, we demonstrated that GTM-1 enhances autophagy clearance and reverses the downregulation of autophagy flux by thapsigargin and asparagine. Furthermore, administration of GTM-1 attenuated Aβ pathology and ameliorated cognitive deficits in AD mice.

Show MeSH
Related in: MedlinePlus