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Synergic prodegradative activity of Bicalutamide and trehalose on the mutant androgen receptor responsible for spinal and bulbar muscular atrophy.

Giorgetti E, Rusmini P, Crippa V, Cristofani R, Boncoraglio A, Cicardi ME, Galbiati M, Poletti A - Hum. Mol. Genet. (2014)

Bottom Line: Thus, (i) prevention of ARpolyQ nuclear localization, combined with (ii) an increased ARpolyQ cytoplasmic clearance, should reduce its detrimental activity.Using the antiandrogen Bicalutamide (Casodex(®)), which slows down AR activation and nuclear translocation, and the disaccharide trehalose, an autophagy activator, we found that, in motoneurons, the two compounds together reduced ARpolyQ insoluble forms with higher efficiency than that obtained with single treatments.Collectively, these data suggest that the combinatory use of Bicalutamide and trehalose is a novel approach to facilitate ARpolyQ clearance that has to be tested in other cell types target of SBMA (i.e. muscle cells) and in vivo in animal models of SBMA.

View Article: PubMed Central - PubMed

Affiliation: Sezione di Biomedicina ed Endocrinologia, Dipartimento di Scienze Farmacologiche e Biomolecolari (DiSFeB), Centro di Eccellenza sulle Malattie Neurodegenerative, Università degli Studi di Milano, Milano 20133, Italy Centro InterUniversitario sulle Malattie Neurodegenerative, Università degli Studi di Firenze, Genova e Roma Tor Vergata, Milano 20133, Italy Department of Pathology, University of Michigan, Ann Arbor, MI 48109, USA and.

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Autophagy activation is mediated by trehalose but not by Bicalutamide. (A) Western blot analysis on mock-transfected NSC34 cells and NSC34 cells transiently transfected with plasmids encoding AR.Q23 or AR.Q46 treated with 10 nm trehalose or 100 nm Bicalutamide (Cas) for 48 h. Two well-known markers of the autophagic pathway (SQSTM1/p62 and LC3-II) were used to compare the activation of autophagy mediated by trehalose or Bicalutamide. GAPDH was used to normalize protein loading. (B) HRFM analysis (63× magnification) on NSC34 cells shows the subcellular distribution and the endogenous expression levels of LC3 and SQSTM1/p62 after 100 nm Bicalutamide (upper panel) or 100 mm trehalose (lower panel) treatments for 48 h, in the presence of mutant AR. Scale bar = 10 μm.
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DDU419F2: Autophagy activation is mediated by trehalose but not by Bicalutamide. (A) Western blot analysis on mock-transfected NSC34 cells and NSC34 cells transiently transfected with plasmids encoding AR.Q23 or AR.Q46 treated with 10 nm trehalose or 100 nm Bicalutamide (Cas) for 48 h. Two well-known markers of the autophagic pathway (SQSTM1/p62 and LC3-II) were used to compare the activation of autophagy mediated by trehalose or Bicalutamide. GAPDH was used to normalize protein loading. (B) HRFM analysis (63× magnification) on NSC34 cells shows the subcellular distribution and the endogenous expression levels of LC3 and SQSTM1/p62 after 100 nm Bicalutamide (upper panel) or 100 mm trehalose (lower panel) treatments for 48 h, in the presence of mutant AR. Scale bar = 10 μm.

Mentions: WB analysis in Figure 2A shows that SQSTM1/p62 levels remains unchanged after Bicalutamide treatment, but are increased by trehalose both in wild-type (wt; AR.Q23) and in ARpolyQ (AR.Q46) expressing cells. Similarly, Bicalutamide has no effect on LC3-I to LC3-II conversion, whereas trehalose stimulates the formation of large amounts of LC3-II form. These data were confirmed in IF microscopy (Fig. 2B), where it is noteworthy that SQSTM1/p62 and LC3 protein expression levels were greatly increased after trehalose treatment, but not by Bicalutamide.Figure 2.


Synergic prodegradative activity of Bicalutamide and trehalose on the mutant androgen receptor responsible for spinal and bulbar muscular atrophy.

Giorgetti E, Rusmini P, Crippa V, Cristofani R, Boncoraglio A, Cicardi ME, Galbiati M, Poletti A - Hum. Mol. Genet. (2014)

Autophagy activation is mediated by trehalose but not by Bicalutamide. (A) Western blot analysis on mock-transfected NSC34 cells and NSC34 cells transiently transfected with plasmids encoding AR.Q23 or AR.Q46 treated with 10 nm trehalose or 100 nm Bicalutamide (Cas) for 48 h. Two well-known markers of the autophagic pathway (SQSTM1/p62 and LC3-II) were used to compare the activation of autophagy mediated by trehalose or Bicalutamide. GAPDH was used to normalize protein loading. (B) HRFM analysis (63× magnification) on NSC34 cells shows the subcellular distribution and the endogenous expression levels of LC3 and SQSTM1/p62 after 100 nm Bicalutamide (upper panel) or 100 mm trehalose (lower panel) treatments for 48 h, in the presence of mutant AR. Scale bar = 10 μm.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4262493&req=5

DDU419F2: Autophagy activation is mediated by trehalose but not by Bicalutamide. (A) Western blot analysis on mock-transfected NSC34 cells and NSC34 cells transiently transfected with plasmids encoding AR.Q23 or AR.Q46 treated with 10 nm trehalose or 100 nm Bicalutamide (Cas) for 48 h. Two well-known markers of the autophagic pathway (SQSTM1/p62 and LC3-II) were used to compare the activation of autophagy mediated by trehalose or Bicalutamide. GAPDH was used to normalize protein loading. (B) HRFM analysis (63× magnification) on NSC34 cells shows the subcellular distribution and the endogenous expression levels of LC3 and SQSTM1/p62 after 100 nm Bicalutamide (upper panel) or 100 mm trehalose (lower panel) treatments for 48 h, in the presence of mutant AR. Scale bar = 10 μm.
Mentions: WB analysis in Figure 2A shows that SQSTM1/p62 levels remains unchanged after Bicalutamide treatment, but are increased by trehalose both in wild-type (wt; AR.Q23) and in ARpolyQ (AR.Q46) expressing cells. Similarly, Bicalutamide has no effect on LC3-I to LC3-II conversion, whereas trehalose stimulates the formation of large amounts of LC3-II form. These data were confirmed in IF microscopy (Fig. 2B), where it is noteworthy that SQSTM1/p62 and LC3 protein expression levels were greatly increased after trehalose treatment, but not by Bicalutamide.Figure 2.

Bottom Line: Thus, (i) prevention of ARpolyQ nuclear localization, combined with (ii) an increased ARpolyQ cytoplasmic clearance, should reduce its detrimental activity.Using the antiandrogen Bicalutamide (Casodex(®)), which slows down AR activation and nuclear translocation, and the disaccharide trehalose, an autophagy activator, we found that, in motoneurons, the two compounds together reduced ARpolyQ insoluble forms with higher efficiency than that obtained with single treatments.Collectively, these data suggest that the combinatory use of Bicalutamide and trehalose is a novel approach to facilitate ARpolyQ clearance that has to be tested in other cell types target of SBMA (i.e. muscle cells) and in vivo in animal models of SBMA.

View Article: PubMed Central - PubMed

Affiliation: Sezione di Biomedicina ed Endocrinologia, Dipartimento di Scienze Farmacologiche e Biomolecolari (DiSFeB), Centro di Eccellenza sulle Malattie Neurodegenerative, Università degli Studi di Milano, Milano 20133, Italy Centro InterUniversitario sulle Malattie Neurodegenerative, Università degli Studi di Firenze, Genova e Roma Tor Vergata, Milano 20133, Italy Department of Pathology, University of Michigan, Ann Arbor, MI 48109, USA and.

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Related in: MedlinePlus