A New Glucocerebrosidase Chaperone Reduces  -Synuclein and Glycolipid Levels in iPSC-Derived Dopaminergic Neurons from Patients with Gaucher Disease and Parkinsonism

Abstract

Among the known genetic risk factors for Parkinson disease, mutations in GBA1, the gene responsible for the lysosomal disorder Gaucher disease, are the most common. This genetic link has directed attention to the role of the lysosome in the pathogenesis of parkinsonism. To study how glucocerebrosidase impacts parkinsonism and to evaluate new therapeutics, we generated induced human pluripotent stem cells from four patients with Type 1 (non-neuronopathic) Gaucher disease, two with and two without parkinsonism, and one patient with Type 2 (acute neuronopathic) Gaucher disease, and differentiated them into macrophages and dopaminergic neurons. These cells exhibited decreased glucocerebrosidase activity and stored the glycolipid substrates glucosylceramide and glucosylsphingosine, demonstrating their similarity to patients with Gaucher disease. Dopaminergic neurons from patients with Type 2 and Type 1 Gaucher disease with parkinsonism had reduced dopamine storage and dopamine transporter reuptake. Levels of ␣-synuclein, a protein present as aggregates in Parkinson disease and related synucleinopathies, were selectively elevated in neurons from the patients with parkinsonism or Type 2 Gaucher disease. The cells were then treated with NCGC607, a small-molecule noninhibitory chaperone of glucocerebrosidase identified by high-throughput screening and medicinal chemistry structure optimization. This compound successfully chaperoned the mutant enzyme, restored glucocerebrosidase activity and protein levels, and reduced glycolipid storage in both iPSC-derived macrophages and dopaminergic neurons, indicating its potential for treating neuronopathic Gaucher disease. In addition, NCGC607 reduced ␣-synuclein levels in dopaminergic neurons from the patients with parkinsonism, suggesting that noninhibitory small-molecule chaperones of glucocerebrosidase may prove useful for the treatment of Parkinson disease. Key words: ␣-synuclein; dopaminergic neurons; glucocerebrosidase; induced pluripotent stem cells; parkinsonism; pharmacological chaperone Significance StatementBecause GBA1 mutations are the most common genetic risk factor for Parkinson disease, dopaminergic neurons were generated from iPSC lines derived from patients with Gaucher disease with and without parkinsonism. These cells exhibit deficient enzymatic activity, reduced lysosomal glucocerebrosidase levels, and storage of glucosylceramide and glucosylsphingosine. Lines generated from the patients with parkinsonism demonstrated elevated levels of ␣-synuclein. To reverse the observed phenotype, the neurons were treated with a novel noninhibitory glucocerebrosidase chaperone, which successfully restored glucocerebrosidase activity and protein levels and reduced glycolipid storage. In addition, the small-molecule chaperone reduced ␣-synuclein levels in dopaminergic neurons, indicating that chaperoning glucocerebrosidase to the lysosome may provide a novel therapeutic strategy for both Parkinson disease and neuronopathic forms of Gaucher disease.

The Complicated Relationship between Gaucher Disease and Parkinsonism: Insights from a Rare Disease

“…Aged neurons from the subjects with PD recapitulate the patient phenotype by displaying increased α-syn levels. This work also demonstrated that iPSC-dopaminergic neurons from patients with type 2 GD, the most severe acute neuronopathic form, also show increased α-syn accumulation (Aflaki et al, 2016). …”

“…It was also found that infants with type 2 GD disease who often have extremely low residual GCase activity exhibit increased neuronal α-syn levels (Aflaki et al, 2016; Mazzulli et al, 2011), although no α-syn pathology is noted at autopsy (Berger-Sieczkowski et al, 2016). …”

“…This property renders them attractive therapeutic candidates for the treatment of synucleinopathies. Two very recent studies on iPSC-derived human midbrain dopaminergic neurons from patients with PD, GD and PD, and GD types 1 and 2 showed that treatment with the non-inhibitory chaperones NCGC758 and NCGC607 reduced substrate accumulation, increased lysosomal activity, enhanced GCase translocation to lysosomes, and reversed α-syn accumulation and downstream toxicity (Aflaki et al, 2016; Mazzulli et al, 2016b). Although such non-inhibitory compounds have not yet been evaluated in animal models, these studies showcase the relevance of appropriate cell-based models in evaluating modulators of GCase as a novel therapy for the synucleinopathies.…”

Glucocerebrosidase Mutations in Parkinson Disease

“…A number of investigations into the use of gene therapy to augment GCase levels in GBAmutation carrying models and models of PD, through adeno-associated virus-mediated (AAV-mediated) expression of GCase in the CNS via cerebral injections has found hugely beneficial effects [88,89], which serves as an excellent proof of concept for the use of GCase activity augmentation in the reduction of alpha-synuclein accumulation and transmission [89,90], and neurodegeneration in PD models [88], which will be an avenue to pursue further with a view to translation to human PD patients.…”

Estudo dos efeitos da superexpressão da alfa-sinucleína sobre o tráfego mitocondrial e autofagia em leveduras, células SH-SY5Y e neurônios dopaminérgicos derivados de hiPSC de pacientes com doença de Parkinson

“…Initially, it was proposed that the proteasome degraded alpha-synuclein. However, recent studies showed that alpha-synuclein is preferentially degraded via lysosomes, especially with protein overexpression (Aflaki et al, 2016;Mak et al, 2010).…”