17-AAG Induces Cytoplasmic α-Synuclein Aggregate Clearance by Induction of Autophagy

Abstract

BackgroundThe accumulation and aggregation of α-synuclein in nerve cells and glia are characteristic features of a number of neurodegenerative diseases termed synucleinopathies. α-Synuclein is a highly soluble protein which in a nucleation dependent process is capable of self-aggregation. The causes underlying aggregate formation are not yet understood, impairment of the proteolytic degradation systems might be involved.Methodology/Principal FindingsIn the present study the possible aggregate clearing effects of the geldanamycin analogue 17-AAG (17-(Allylamino)-17-demethoxygeldanamycin) was investigated. Towards this, an oligodendroglial cell line (OLN-93 cells), stably expressing human α-synuclein (A53T mutation) was used. In these cells small punctate aggregates, not staining with thioflavine S, representing prefibrillary aggregates, occur characteristically. Our data demonstrate that 17-AAG attenuated the formation of α-synuclein aggregates by stimulating macroautophagy. By blocking the lysosomal compartment with NH4Cl the aggregate clearing effects of 17-AAG were abolished and α-synuclein deposits were enlarged. Analysis of LC3-II immunoreactivity, which is an indicator of autophagosome formation, further revealed that 17-AAG led to the recruitment of LC3-II and to the formation of LC3 positive puncta. This effect was also observed in cultured oligodendrocytes derived from the brains of newborn rats. Inhibition of macroautophagy by 3-methyladenine prevented 17-AAG induced occurrence of LC3 positive puncta as well as the removal of α-synuclein aggregates in OLN-A53T cells.ConclusionsOur data demonstrate for the first time that 17-AAG not only causes the upregulation of heat shock proteins, but also is an effective inducer of the autophagic pathway by which α-synuclein can be removed. Hence geldanamycin derivatives may provide a means to modulate autophagy in neural cells, thereby ameliorating pathogenic aggregate formation and protecting the cells during disease and aging.

Chronic Treatment with 17-DMAG Improves Balance and Coordination in A New Mouse Model of Machado-Joseph Disease

“…Since it was previously shown that 17-AAG and its analogue used in this study-17-DMAG-are able to activate the autophagy process in different model systems [35][36][37], and given that our results suggested degradation of ataxin-3 was occurring, we assessed known autophagy markers in 17-DMAG-treated animals. Our results suggest that autophagy is activated by the drug specifically in diseased animals.…”

“…These results suggest that HSR induction by chronic administration of 17-DMAG at 25 mg/kg may be compromised in the disease state, particularly at later stages. Since it was previously shown that 17-AAG and its analogue 17-DMAG are able to activate macroautophagy in different model systems [35][36][37], we also assessed activation of this pathway. We observed an increase in the beclin-1 protein levels in the brainstem of 16-and 30-week-old mice and of the LC3-II/LC3-I ratio at 30 weeks of age, after chronic 17-DMAG treatment (p=0.008, p=0.017, respectively) in transgenic mice but not in controls, confirming the specificity of this activation (Fig.…”

A Novel Hsp90 Inhibitor Activates Compensatory Heat Shock Protein Responses and Autophagy and Alleviates Mutant A53T α-Synuclein Toxicity

“…However, the failure of these candidate molecules in ameliorating dopaminergic neurodegeneration indicated that the compensatory upregulation of Hsp70 by itself may be insufficient to protect against a-Syn toxicity (Zourlidou et al, 2004;Shimshek et al, 2010). The BQA class of Hsp90 inhibitors, including GA and 17-AAG, has been found to be effective in inducing both multiple HSPs and autophagy (Finn et al, 2005;Riedel et al, 2010;Watanabe et al, 2010). However, the protective effects of BQAs are accompanied by off-target toxicities, limiting their use in neurodegenerative diseases.…”

“…Pharmacologic inhibition of Hsp90 has been shown to be useful in neurodegenerative disease models (Gallo, 2006;Luo et al, 2010). There are at least three potential protective mechanisms associated with inhibition of Hsp90: 1) Hsp90 inhibition leads to activation of heat shock factor 1, which further induces a range of small heat shock proteins (HSPs; including Hsp70 and Hsp27) that assist in appropriate folding of misfolded proteins (Dimant et al, 2012); 2) inhibition of Hsp90 leads to the degradation of mutant neuronal client proteins such as polyglutamineexpanded mutant androgen receptor and leucine-rich repeat kinase 2, which reduces the load of toxic Hsp90 client proteins (Waza et al, 2005;Tokui et al, 2009;Taipale et al, 2010;Baldo et al, 2012); and 3) Hsp90 inhibitors including 17-(allylamino)-17-demethoxygeldanamycin (17-AAG) have been shown to induce protective autophagy that led to degradation of mutant and misfolded protein aggregates (Riedel et al, 2010). The benzoquinone ansamycin (BQA) Hsp90 inhibitors geldanamycin (GA), 17-AAG, and 17-(dimethylaminoethylamino)-17-demethoxygeldanamycin (17-DMAG) have been shown to induce a robust compensatory heat shock response and amelioration of protein aggregation and toxicity.…”

Aberrations in Proteostasis Orchestrate The Genotypic and Phenotypic Changes in Aging

“…Small chemical activators of heat shock transcription factor 1 (HSF1), such as geldanamycin induce multiple endogenous molecular chaperones and have already shown to be effective in limiting protein aggregation models including drosophila model of polyQ disease (Fujikake et al, 2008;Riedel et al, 2010).…”