A Protein Aggregation Inhibitor, Leuco-Methylthioninium Bis(Hydromethanesulfonate), Decreases α-Synuclein Inclusions in a Transgenic Mouse Model of Synucleinopathy

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Abstract

α-Synuclein (α-Syn) aggregation is a pathological feature of synucleinopathies, neurodegenerative disorders that include Parkinson's disease (PD). We have tested whether N,N,N',N'-tetramethyl-10H-phenothiazine-3,7-diaminium bis(hydromethanesulfonate) (leuco-methylthioninium bis(hydromethanesulfonate); LMTM), a tau aggregation inhibitor, affects α-Syn aggregation in vitro and in vivo. Both cellular and transgenic models in which the expression of full-length human α-Syn (h-α-Syn) fused with a signal sequence peptide to promote α-Syn aggregation were used. Aggregated α-Syn was observed following differentiation of N1E-115 neuroblastoma cells transfected with h-α-Syn. The appearance of aggregated α-Syn was inhibited by LMTM, with an EC50 of 1.1 μM, with minimal effect on h-α-Syn mRNA levels being observed. Two independent lines of mice (L58 and L62) transgenic for the same fusion protein accumulated neuronal h-α-Syn that, with aging, developed into fibrillary inclusions characterized by both resistance to proteinase K (PK)-cleavage and their ability to bind thiazin red. There was a significant decrease in α-Syn-positive neurons in multiple brain regions following oral treatment of male and female mice with LMTM administered daily for 6 weeks at 5 and 15 mg MT/kg. The early aggregates of α-Syn and the late-stage fibrillar inclusions were both susceptible to inhibition by LMTM, a treatment that also resulted in the rescue of movement and anxiety-related traits in these mice. The results suggest that LMTM may provide a potential disease modification therapy in PD and other synucleinopathies through the inhibition of α-Syn aggregation.

Original languageEnglish
Article number447
Number of pages15
JournalFrontiers in Molecular Neuroscience
Volume10
DOIs
Publication statusPublished - 10 Jan 2018

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Synucleins
Transgenic Mice
Proteins
Parkinson Disease
Endopeptidase K
Aptitude
Protein Sorting Signals
Neuroblastoma
Neurodegenerative Diseases
Anxiety
Neurons
Messenger RNA
Brain

Keywords

  • Journal Article
  • aggregation inhibitor
  • leucomethylthioninium
  • mouse model
  • Parkinson’s disease
  • α-Synuclein
  • synucleinopathy

Cite this

@article{b0597bf533b0487482edb0faf565b61b,
title = "A Protein Aggregation Inhibitor, Leuco-Methylthioninium Bis(Hydromethanesulfonate), Decreases α-Synuclein Inclusions in a Transgenic Mouse Model of Synucleinopathy",
abstract = "α-Synuclein (α-Syn) aggregation is a pathological feature of synucleinopathies, neurodegenerative disorders that include Parkinson's disease (PD). We have tested whether N,N,N',N'-tetramethyl-10H-phenothiazine-3,7-diaminium bis(hydromethanesulfonate) (leuco-methylthioninium bis(hydromethanesulfonate); LMTM), a tau aggregation inhibitor, affects α-Syn aggregation in vitro and in vivo. Both cellular and transgenic models in which the expression of full-length human α-Syn (h-α-Syn) fused with a signal sequence peptide to promote α-Syn aggregation were used. Aggregated α-Syn was observed following differentiation of N1E-115 neuroblastoma cells transfected with h-α-Syn. The appearance of aggregated α-Syn was inhibited by LMTM, with an EC50 of 1.1 μM, with minimal effect on h-α-Syn mRNA levels being observed. Two independent lines of mice (L58 and L62) transgenic for the same fusion protein accumulated neuronal h-α-Syn that, with aging, developed into fibrillary inclusions characterized by both resistance to proteinase K (PK)-cleavage and their ability to bind thiazin red. There was a significant decrease in α-Syn-positive neurons in multiple brain regions following oral treatment of male and female mice with LMTM administered daily for 6 weeks at 5 and 15 mg MT/kg. The early aggregates of α-Syn and the late-stage fibrillar inclusions were both susceptible to inhibition by LMTM, a treatment that also resulted in the rescue of movement and anxiety-related traits in these mice. The results suggest that LMTM may provide a potential disease modification therapy in PD and other synucleinopathies through the inhibition of α-Syn aggregation.",
keywords = "Journal Article, aggregation inhibitor, leucomethylthioninium, mouse model, Parkinson’s disease, α-Synuclein, synucleinopathy",
author = "Karima Schwab and Silke Frahm and David Horsley and Rickard, {Janet E} and Valeria Melis and Goatman, {Elizabeth A} and Mandy Magbagbeolu and Morag Douglas and Leith, {Michael G} and Baddeley, {Thomas C.} and Storey, {John M D} and Gernot Riedel and Wischik, {Claude M} and Harrington, {Charles R} and Franz Theuring",
note = "The authors acknowledge Heide Lueck for excellent technical assistance and for maintenance of animals. AMS Biotechnology generated monoclonal antibodies from recombinant α-Syn prepared by JR.",
year = "2018",
month = "1",
day = "10",
doi = "10.3389/fnmol.2017.00447",
language = "English",
volume = "10",
journal = "Frontiers in Molecular Neuroscience",
issn = "1662-5099",
publisher = "Frontiers Media S.A.",

}

TY - JOUR

T1 - A Protein Aggregation Inhibitor, Leuco-Methylthioninium Bis(Hydromethanesulfonate), Decreases α-Synuclein Inclusions in a Transgenic Mouse Model of Synucleinopathy

AU - Schwab, Karima

AU - Frahm, Silke

AU - Horsley, David

AU - Rickard, Janet E

AU - Melis, Valeria

AU - Goatman, Elizabeth A

AU - Magbagbeolu, Mandy

AU - Douglas, Morag

AU - Leith, Michael G

AU - Baddeley, Thomas C.

AU - Storey, John M D

AU - Riedel, Gernot

AU - Wischik, Claude M

AU - Harrington, Charles R

AU - Theuring, Franz

N1 - The authors acknowledge Heide Lueck for excellent technical assistance and for maintenance of animals. AMS Biotechnology generated monoclonal antibodies from recombinant α-Syn prepared by JR.

PY - 2018/1/10

Y1 - 2018/1/10

N2 - α-Synuclein (α-Syn) aggregation is a pathological feature of synucleinopathies, neurodegenerative disorders that include Parkinson's disease (PD). We have tested whether N,N,N',N'-tetramethyl-10H-phenothiazine-3,7-diaminium bis(hydromethanesulfonate) (leuco-methylthioninium bis(hydromethanesulfonate); LMTM), a tau aggregation inhibitor, affects α-Syn aggregation in vitro and in vivo. Both cellular and transgenic models in which the expression of full-length human α-Syn (h-α-Syn) fused with a signal sequence peptide to promote α-Syn aggregation were used. Aggregated α-Syn was observed following differentiation of N1E-115 neuroblastoma cells transfected with h-α-Syn. The appearance of aggregated α-Syn was inhibited by LMTM, with an EC50 of 1.1 μM, with minimal effect on h-α-Syn mRNA levels being observed. Two independent lines of mice (L58 and L62) transgenic for the same fusion protein accumulated neuronal h-α-Syn that, with aging, developed into fibrillary inclusions characterized by both resistance to proteinase K (PK)-cleavage and their ability to bind thiazin red. There was a significant decrease in α-Syn-positive neurons in multiple brain regions following oral treatment of male and female mice with LMTM administered daily for 6 weeks at 5 and 15 mg MT/kg. The early aggregates of α-Syn and the late-stage fibrillar inclusions were both susceptible to inhibition by LMTM, a treatment that also resulted in the rescue of movement and anxiety-related traits in these mice. The results suggest that LMTM may provide a potential disease modification therapy in PD and other synucleinopathies through the inhibition of α-Syn aggregation.

AB - α-Synuclein (α-Syn) aggregation is a pathological feature of synucleinopathies, neurodegenerative disorders that include Parkinson's disease (PD). We have tested whether N,N,N',N'-tetramethyl-10H-phenothiazine-3,7-diaminium bis(hydromethanesulfonate) (leuco-methylthioninium bis(hydromethanesulfonate); LMTM), a tau aggregation inhibitor, affects α-Syn aggregation in vitro and in vivo. Both cellular and transgenic models in which the expression of full-length human α-Syn (h-α-Syn) fused with a signal sequence peptide to promote α-Syn aggregation were used. Aggregated α-Syn was observed following differentiation of N1E-115 neuroblastoma cells transfected with h-α-Syn. The appearance of aggregated α-Syn was inhibited by LMTM, with an EC50 of 1.1 μM, with minimal effect on h-α-Syn mRNA levels being observed. Two independent lines of mice (L58 and L62) transgenic for the same fusion protein accumulated neuronal h-α-Syn that, with aging, developed into fibrillary inclusions characterized by both resistance to proteinase K (PK)-cleavage and their ability to bind thiazin red. There was a significant decrease in α-Syn-positive neurons in multiple brain regions following oral treatment of male and female mice with LMTM administered daily for 6 weeks at 5 and 15 mg MT/kg. The early aggregates of α-Syn and the late-stage fibrillar inclusions were both susceptible to inhibition by LMTM, a treatment that also resulted in the rescue of movement and anxiety-related traits in these mice. The results suggest that LMTM may provide a potential disease modification therapy in PD and other synucleinopathies through the inhibition of α-Syn aggregation.

KW - Journal Article

KW - aggregation inhibitor

KW - leucomethylthioninium

KW - mouse model

KW - Parkinson’s disease

KW - α-Synuclein

KW - synucleinopathy

U2 - 10.3389/fnmol.2017.00447

DO - 10.3389/fnmol.2017.00447

M3 - Article

VL - 10

JO - Frontiers in Molecular Neuroscience

JF - Frontiers in Molecular Neuroscience

SN - 1662-5099

M1 - 447

ER -