Abstract Body

Aims

Parkinson’s disease (PD)-affected brains show endoplasmic reticulum (ER) stress and mitophagic dysfunctions. How these processes are altered and whether they are directly linked remain a matter of questions. XBP1 is a transcription factor activated upon ER stress after unconventional splicing by the nuclease IREa thereby yielding XBP1S. PINK1 is a kinase considered as the sensor of mitochondrial physiology and a master gatekeeper of mitophagy process. Our work aims at delineating the putative link between XBP1S and PINK1 that could explain ER stress alteration and mitophagic defects in Parkinsons disease.

Methods

We have used cell biology, molecular biology and pharmacological approaches to unravel a forward loop regulation between XBP1 and PINK1 in various neuronal models and in vivo. Extensive ex-vivo and in vivo mitophagy analysis by biochemical approaches and evaluation of mitochondrial function (mitochondrial potential, Δψmit) and morphology were assessed by combined procedures including, flow cytometry, live imaging analysis and electronic microcopy.

Results

We show that XBP1S transactivates PINK1 in human cells, primary cultured neurons and mice brain, and triggers a pro-mitophagic phenotype that is fully dependent of endogenous PINK1. We also unravel a PINK1-dependent phosphorylation of XBP1S that conditions its nuclear localization and thereby, governs its transcriptional activity. PINK1-induced XBP1S phosphorylation occurs at residues reminiscent of those phosphorylated in substantia nigra of PD-affected brains.

Conclusions

Overall, our study delineates for the first time a functional forward loop between XBP1S and PINK1 governing a molecular link between ER-stress and mitophagy that could be disrupted in PD condition

Aims

Parkinson’s disease (PD) has useful symptomatic treatments that do not slow the neurodegenerative process, and no significant disease-modifying treatments are approved. A key therapeutic target in PD is α-synuclein (αS), which is both genetically implicated and accumulates in Lewy bodies rich in vesicles and other lipid membranes. Reestablishing αS-homeostasis is a central goal in PD. Based on previous lipidomic analyses, we conducted a mouse-trial of a stearoyl-CoA desaturase (SCD) inhibitor, '5b', that prevented αS-positive vesicular inclusions and cytotoxicity in cultured human neurons.

Methods

Oral dosing and brain activity of 5b were established in non-tg mice. 5b in drinking water was given to mice expressing WT hu αS or an amplified familial PD αS mutation [E35K+E46K+E61K (‘3K’)] beginning near the onset of nigral and cortical neurodegeneration and the robust PD-like motor syndrome in 3K. Motor phenotypes, brain cytopathology and SCD-related lipid changes were quantified in 5b- vs. placebo-treated mice. Outcomes were compared to effects of crossing 3K to SCD1^-/- mice.

Results

5b treatment reduced αS hyperphosphorylation in E46K-expresing human neurons, in 3K neural cultures and in both WT hu and 3K αS mice. 5b prevented subtle gait deficits in WT hu αS mice and the PD-like resting tremor and progressive motor decline of 3K mice. 5b also increased αS tetramers and reduced PK-resistant lipid-rich aggregates. Similar benefits accrued from genetically deleting one SCD allele, providing target validation.

Conclusions

Prolonged reduction of brain SCD activity prevented PD-like neuropathology in multiple PD models. Thus, an orally available SCD inhibitor potently ameliorates PD phenotypes, positioning this approach to treat human α-synucleinopathies.

Aims

To evaluate YTX-7739, a clinical stage stearoyl-CoA desaturase (SCD) inhibitor, in the fPD E46K-amplified “3K” alpha-synuclein mouse model that recapitulates key motor and pathological features of Parkinson’s disease.

Methods

YTX-7739 was formulated in food at a dose shown to reduce the fatty acid desaturation index, a biomarker of SCD activity. Mice expressing WT or 3K mutant alpha-synuclein were fed YTX-7739 ad libitum from age 2 to 6 months. Motor behavior, biochemistry and histopathology were assessed. Compound concentrations and fatty acid profiles were quantified.

Results

3K and WT alpha-synuclein mice receiving YTX-7739 had an 80% reduction in the brain C16 desaturation index (16:1/16:0) at the end of treatment. This decrease was associated with improved motor function in pole-climbing and rotarod tests. Postmortem biochemical and immunohistochemical analyses showed a YTX-7739-dependent reduction in alpha-synuclein pathology, including increased alpha-synuclein tetramer:monomer ratio, reduced levels of buffer-insoluble alpha-synuclein, and decreased phospho-S129 and PK-resistant alpha-synuclein. YTX-7739 also prevented the loss of TH-positive fibers and reduced the presence of lipid droplets in neurons. The effects of YTX-7739 on alpha-synuclein pathologies in the mouse model were recapitulated in patient-derived iPSC neurons, demonstrating the consistency of effect from mouse to human cells.

Conclusions

SCD inhibition by YTX-7739 improved motor function and mitigated pathology in an alpha-synuclein mouse model that shows behavioral and pathological features of Parkinson’s disease. These data highlight the critical role of lipid biology, especially saturated fatty acids, in mediating alpha-synuclein pathobiology and supports the on-going clinical evaluation of YTX-7739 as a disease-modifying drug for synucleinopathies.

Aims

Parkinson’s disease (PD) is characterized by a progressive loss of nigral dopamine neurons and deposition of fibrillary aggregated α-synuclein in Lewy bodies (LB). We recently described that synapsin III (Syn III), a synaptic phosphoprotein regulating dopamine release in cooperation with α-synuclein, composes LB insoluble fibrils in PD patients brains, where a marked α-synuclein/Syn III neuropathology was detectable by the in situ proximity ligation assay. We thus investigated whether Syn III might constitute a novel therapeutic target for PD by studying its involvement in α-synuclein aggregation and in the associated nigrostriatal synaptic damage/degeneration and motor symptom onset.

Methods

Different PD in vivo models were used and gene silencing as well as drug-mediated Syn III manipulation were exploited.

Results

We found that Syn III ko mice did not develop fibrillary insoluble α-synuclein aggregates and that their nigrostriatal neurons were protected from both synaptic alterations and degeneration resulting from adeno-associated vector-mediated α-synuclein overexpression. Moreover, gene silencing of Syn III in a human α-synuclein transgenic mouse model at pathological stage could revert α-synuclein aggregation, synaptic derangement and motor symptom onset. Finally, we observed that the monoamine inhibitor methylphenidate, owning Syn III-binding ability, induced a Syn III-reliant motor response in the human α-synuclein transgenic mice independently of its dopamine transporter inhibitory action.

Conclusions

This observation provide a rationale for the freezing of gait-counteracting activity of methylphenidate in advanced PD. Our findings indicate that Syn III is as an accessory mediator of α-synuclein aggregation and toxicity as well as a druggable therapeutic target for PD.

Aims

Accumulation of abnormal protein aggregates is a paramount pathological hallmark shared by several, if not all, neurodegenerative diseases. One example is α-synuclein-rich inclusions found in synucleinopathies, which are referred to as Lewy bodies. While their presence is well established, how these aggregates affect neuronal homeostasis remains elusive, in part because we lack the proper tools to undertake such investigations.

Methods

We have therefore set out to tackle this question by developing a unique approach to fully control α-syn aggregation and LBs formation in cell culture and within cerebral tissue using light. We have termed this technique Light-inducible protein aggregation (LIPA) and herein provide the extent of applicability using both in vitro and in vivo contexts.

Results

In living cells and in vivo, our LIPA system allows real-time induction of α-syn inclusion formation with remarkable spatial and temporal resolution. LIPA-induced aggregates auto-perpetuate for several days after transient induction with light, and faithfully mimic several authentic features of LBs.

Conclusions

This system constitutes a new and exceptional tool by which to generate, visualize and dissect the role of LBs in neurodegeneration and as such, identify new genetic and pharmacological inhibitors of α-syn aggregation for the treatment of α-synucleinopathies.

Aims

To understand the function and regulation of aldehyde dehydrogenase 1a1 (ALDH1A1)-positive nigrostriatal dopaminergic neurons (ALDH1A1⁺ nDANs), which display the most profound loss in Parkinson's disease (PD).

Methods

Using custom-built ALDH1A1-CreERT2 knock-in mice, we selectively ablated the ALDH1A1⁺ nDANs and tested for any PD-related motor and non-motor phenotypes. We then determined the connectivity of these neurons and examined the impact of different presynaptic inputs on the function of these neurons. We further performed live imaging of dopamine release in behaving mice to correlate neuron activity with different behavioral modalities.

Results

We presented the first whole-brain circuit map of ALDH1A1⁺ nDANs and revealed an essential physiological function of these neurons in regulating the vigor of movement during the acquisition of motor skills. We found that the ALDH1A1⁺ nDAN-ablated mice moved less often at high walking-speed and completely lost the ability to improve the vigor of movement during the motor skill learning task. Dopamine replacement therapy, routinely used to treat PD patients was effective to restore walking speed, but it failed to rescue motor learning impairments. Live imaging studies further revealed a close correlation of dopamine release with the motor performance during the acquisition phase of motor learning.

Conclusions

This study underscores the importance of intact brain circuit, which allows the ALDH1A1⁺ nDAN to integrate diverse presynaptic inputs and produce timely and dynamic dopamine release in the motor learning process.

Aims

Parkinson’s disease (PD) is characterized by the gradual appearance of intraneuronal Lewy aggregates (Lewy bodies), which are primarily composed of misfolded α-synuclein (α-syn), resulting in cytotoxicity and neural death. Recent studies support the idea of transcellular spread of misfolded α-syn in a prion-like transmission via exosomes secretion and induce pathological aggregates in healthy neurons. Indeed, exosomes derived from brain lysates and cerebrospinal fluid (CSF) of PD patients have been shown to propagate α-syn aggregation in healthy cells. Braak et al hypothesized the olfactory bulb as primary propagation site for the initiation of the disease. In the current study we examined whether aggregated α-syn can actively spread from the nasal cavity to the brain via exosomes and initiate pathological aggregate in the brain.

Methods

Healthy 2-months old C57/Bl mice were given exosomes isolated from patient's CSF, PD or non synucleinopathy disorders, by internasal administration, and three months after treatment, mice were subjected to behavior tests.

Results

CSF-derived exosomes from PD patients induced aggregation of α-syn in a neuroblastoma cell line as can be seen by positive Thioflavins staining. Importantly, internasal delivery of CSF-derived exosomes from PD patients into the nasal cavity of healthy wild type mice induced PD-like symptoms including impairments in motor function, hyposmia and elevated anxiety, compared to Non-PD treated and healthy untreated mice.

Conclusions

Our data suggests that CSF-derived exosomes from PD patients are sufficient for propagating α-syn aggregation in cell culture and initiating PD-like symptoms in healthy mice after internasal delivery.