Inhibition of TDP-43 accumulation by bis(thiosemicarbazonato)-copper complexes.

Sarah J Parker,Katja M Kanninen,Jeffrey R Liddell,Takashi Nonaka,Brett M Paterson,Sinchun Lim,Peter J Crouch,Janine L James,Anthony R White,Masato Hasegawa,Jodi Meyerowitz,Paul S Donnelly,Philipp J Kahle

doi:10.1371/journal.pone.0042277

Abstract

Amyotrophic lateral sclerosis (ALS) is a progressive, fatal, motor neuron disease with no effective long-term treatment options. Recently, TDP-43 has been identified as a key protein in the pathogenesis of some cases of ALS. Although the role of TDP-43 in motor neuron degeneration is not yet known, TDP-43 has been shown to accumulate in RNA stress granules (SGs) in cell models and in spinal cord tissue from ALS patients. The SG association may be an early pathological change to TDP-43 metabolism and as such a potential target for therapeutic intervention. Accumulation of TDP-43 in SGs induced by inhibition of mitochondrial activity can be inhibited by modulation of cellular kinase activity. We have also found that treatment of cells and animal models of neurodegeneration, including an ALS model, with bioavailable bis(thiosemicarbazonato)copperII complexes (CuII(btsc)s) can modulate kinase activity and induce neuroprotective effects. In this study we examined the effect of diacetylbis(-methylthiosemicarbazonato)copperII (CuII(atsm)) and glyoxalbis(-methylthiosemicarbazonato)copperII (CuII(gtsm)) on TDP-43-positive SGs induced in SH-SY5Y cells in culture. We found that the CuII(btsc)s blocked formation of TDP-43-and human antigen R (HuR)-positive SGs induced by paraquat. The CuII(btsc)s protected neurons from paraquat-mediated cell death. These effects were associated with inhibition of ERK phosphorylation. Co-treatment of cultures with either CuII(atsm) or an ERK inhibitor, PD98059 both prevented ERK activation and blocked formation of TDP-43-and HuR-positive SGs. CuII(atsm) treatment or ERK inhibition also prevented abnormal ubiquitin accumulation in paraquat-treated cells suggesting a link between prolonged ERK activation and abnormal ubiquitin metabolism in paraquat stress and inhibition by Cu. Moreover, CuII(atsm) reduced accumulation of C-terminal (219–414) TDP-43 in transfected SH-SY5Y cells. These results demonstrate that CuII(btsc) complexes could potentially be developed as a neuroprotective agent to modulate neuronal kinase function and inhibit TDP-43 aggregation. Further studies in TDP-43 animal models are warranted.

Highlights

Amyotrophic lateral sclerosis (ALS) is a fatal adult-onset motor neuron disease in which death of upper and lower motor neurons leads to progressive brain and spinal cord deterioration
As CuII(atsm) was found to inhibit extracellular signal-regulated kinase (ERK) phosphorylation in SHSY5Y cells treated with paraquat, we compared the effect of CuII(atsm) and an ERK inhibitor on stress granules (SGs) formation. We found that both CuII(atsm) and PD98059 (ERK inhibitor) induced the same inhibition of TAR DNA binding protein 43 (TDP-43) and human antigen R (HuR)-positive SGs when co-treated with paraquat (compare Figure 5G–I (CuII(atsm) with Figure 5J–L (PD98059)) and Figure 5P
We investigated the ability of CuII(btsc) Cucomplexes to inhibit stress-mediated TDP-43-positive SGs in neuronal cell culture

Summary

Introduction

Amyotrophic lateral sclerosis (ALS) is a fatal adult-onset motor neuron disease in which death of upper and lower motor neurons leads to progressive brain and spinal cord deterioration. Studies into ALS identified familial mutations in a small percentage of patients. Extensive cell and animal-based studies have focused on SOD1 biology in ALS [2,3], these mutations only account for around 20% of familial ALS cases and approximately 2–3% of all ALS patients [4,5]. TDP-43-positive inclusions have been identified in a number of neurodegenerative diseases. In these cases, the TDP43 identification is referred to as a secondary TDP-43 proteinopathy [8]. While the role of abnormal TDP-43 accumulation in both primary and secondary TDP-43 proteinopathies is not yet fully understood, the identification of TDP-43 mutations associated with ALS and FTD has provided clear evidence that altered

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