Using artificial intelligence to identify drugs for repurposing to treat l-DOPA-induced dyskinesia.

Repurposing regulatory agency-approved molecules, with proven safety in humans, is an attractive option for developing new treatments for disease. We identified and assessed the efficacy of 3 drugs predicted by an in silico screen as having the potential to treat l-DOPA-induced dyskinesia (LID) in Parkinson's disease. We analysed ∼1.3 million Medline abstracts using natural language processing and ranked 3539 existing drugs based on predicted ability to reduce LID. 3 drugs from the top 5% of the 3539 candidates; lorcaserin, acamprosate and ganaxolone, were prioritized for preclinical testing based on i) having a novel mechanism of action, ii) having not been previously validated for the treatment of LID, iii) being blood-brain-barrier penetrant and orally bioavailable and iv) being clinical trial ready. We assessed the efficacy of acamprosate, ganaxolone and lorcaserin in a rodent model of l-DOPA-induced hyperactivity, with lorcaserin affording a 58% reduction in rotational asymmetry (P < 0.05) compared to vehicle. Acamprosate and ganaxolone failed to demonstrate efficacy. Lorcaserin, a 5HT2C agonist, was then further tested in MPTP lesioned dyskinetic macaques where it afforded an 82% reduction in LID (P < 0.05), unfortunately accompanied by a significant increase in parkinsonian disability. In conclusion, although our data do not support the repurposing of lorcaserin, acamprosate or ganaxolone per se for LID, we demonstrate value of an in silico approach to identify candidate molecules which, in combination with an in vivo screen, can facilitate clinical development decisions. The present study adds to a growing literature in support of this paradigm shifting approach in the repurposing pipeline.

Prevalence and Distribution of Lewy Pathology in a Homeless Population.

BACKGROUND: The homeless population experience significant inequalities in health, and there is an increasing appreciation of the potential of lifestyle factors in the development of neurodegenerative diseases, including Parkinson's disease. We performed a study on the prevalence and distribution of pathological alpha-synuclein deposition throughout the central and peripheral nervous systems in a homeless population. METHODS: Forty-four homeless individuals consecutively available for autopsy were recruited. Immunohistochemistry was performed using 5G4 antibody recognizing disease-associated forms of alpha-synuclein, complemented by phospho-synuclein antibody on autopsy tissues collected from 18 regions of the brain and spinal cord, as well as the right and left olfactory bulb, the cauda equina, the extramedullary portion of the vagus nerve, and 27 sites of peripheral organs. RESULTS: The study cohort consisted of 38 males and 6 females, median age 58 years (range 32-67). Lewy-related pathology was present in the brains of three male cases. One showed Braak stage 2 (60 years old), and two stage 4 (56 and 59 years old). One of the Braak stage 4 cases had Lewy-related pathology in the spinal cord, the cauda equina, and the extramedullary portion of the vagus nerve. Examination of 27 sites of peripheral organs found that all three cases with Lewy-related pathology present in the brain were devoid of peripheral organ alpha-synuclein pathology. Multiple system-type alpha-synuclein pathology was not found. CONCLUSION: Our study, representing a snapshot of the homeless population that came to autopsy, suggests that alpha-synuclein pathology is prevalent in the homeless supporting further study of this vulnerable population.

4R-Tau seeding activity unravels molecular subtypes in patients with Progressive Supranuclear Palsy.

Progressive Supranuclear palsy (PSP) is a 4-repeat (4-R) tauopathy. We hypothesized that the molecular diversity of tau could explain the heterogeneity seen in PSP disease progression. To test this hypothesis, we performed an extensive biochemical characterisation of the high molecular weight tau species (HMW-Tau) in 20 different brain regions of 25 PSP patients. We found a correlation between the HMW-Tau species and tau seeding capacity in the primary motor cortex, where we confirmed that an elevated 4R-Tau seeding activity correlates with a shorter disease duration. To identify factors that contribute to these differences, we performed proteomic and spatial transcriptomic analysis that revealed key mechanistic pathways, in particular those involving the immune system, that defined patients demonstrating high and low tau seeding capacity. These observations suggest that differences in the tau seeding activity may contribute to the considerable heterogeneity seen in disease progression of patients suffering from PSP.

An AI-guided screen identifies probucol as an enhancer of mitophagy through modulation of lipid droplets.

Failures in mitophagy, a process by which damaged mitochondria are cleared, results in neurodegeneration, while enhancing mitophagy promotes the survival of dopaminergic neurons. Using an artificial intelligence platform, we employed a natural language processing approach to evaluate the semantic similarity of candidate molecules to a set of well-established mitophagy enhancers. Top candidates were screened in a cell-based mitochondrial clearance assay. Probucol, a lipid-lowering drug, was validated across several orthogonal mitophagy assays. In vivo, probucol improved survival, locomotor function, and dopaminergic neuron loss in zebrafish and fly models of mitochondrial damage. Probucol functioned independently of PINK1/Parkin, but its effects on mitophagy and in vivo depended on ABCA1, which negatively regulated mitophagy following mitochondrial damage. Autophagosome and lysosomal markers were elevated by probucol treatment in addition to increased contact between lipid droplets (LDs) and mitochondria. Conversely, LD expansion, which occurs following mitochondrial damage, was suppressed by probucol and probucol-mediated mitophagy enhancement required LDs. Probucol-mediated LD dynamics changes may prime the cell for a more efficient mitophagic response to mitochondrial damage.

Cell-Specific Dysregulation of Iron and Oxygen Homeostasis as a Novel Pathophysiology in PSP.

OBJECTIVE: Progressive supranuclear palsy (PSP) is a 4R-tauopathy showing heterogeneous tau cytopathology commencing in the globus pallidus (GP) and the substantia nigra (SN), regions also associated with age-related iron accumulation. Abnormal iron levels have been extensively associated with tau pathology in neurodegenerative brains, however, its role in PSP pathogenesis remains yet unknown. We perform the first cell type-specific evaluation of PSP iron homeostasis and the closely related oxygen homeostasis, in relation to tau pathology in human postmortem PSP brains. METHODS: In brain regions vulnerable to PSP pathology (GP, SN, and putamen), we visualized iron deposition in tau-affected and unaffected neurons, astroglia, oligodendrocytes, and microglia, using a combination of iron staining with immunolabelling. To further explore molecular pathways underlying our observations, we examined the expression of key iron and oxygen homeostasis mRNA transcripts and proteins. RESULTS: We found astrocytes as the major cell type accumulating iron in the early affected regions of PSP, highly associated with cellular tau pathology. The same regions are affected by dysregulated expression of alpha and beta hemoglobin and neuroglobin showing contrasting patterns. We discovered changes in iron and oxygen homeostasis-related gene expression associated with aging of the brain, and identified dysregulated expression of rare neurodegeneration with brain iron accumulation (NBIA) genes associated with tau pathology to distinguish PSP from the healthy aging brain. INTERPRETATION: We present novel aspects of PSP pathophysiology highlighting an overlap with NBIA pathways. Our findings reveal potential novel targets for therapy development and have implications beyond PSP for other iron-associated neurodegenerative diseases. ANN NEUROL 2023;93:431-445.

Epigenetic Clock Acceleration Is Linked to Age at Onset of Parkinson's Disease.

BACKGROUND: Aging is the strongest risk factor for Parkinson's disease (PD), which is a clinically heterogeneous movement disorder with highly variable age at onset. DNA methylation age (DNAm age) is an epigenetic clock that could reflect biological aging. OBJECTIVES: The aim was to evaluate whether PD age at onset is associated with DNAm-age acceleration (difference between DNAm age and chronological age). METHODS: We used the genome-wide Infinium MethylationEPIC array to assess DNAm age in discovery (n = 96) and replication (n = 182) idiopathic PD cohorts and a unique longitudinal LRRK2 cohort (n = 220) at four time points over a 3-year period, comprising 91 manifesting and 129 nonmanifesting G2019S carriers at baseline. Cox proportional hazard regression and multivariate linear regression were used to evaluate the relation between DNAm-age acceleration and PD age at onset, which was highly variable in manifesting G2019S carriers (36-75 years) and both idiopathic PD cohorts (26-77 and 35-81 years). RESULTS: DNAm-age acceleration remained steady over the 3-year period in most G2019S carriers. It was strongly associated with age at onset in the LRRK2 cohort (P = 2.25 × 10-15 ) and discovery idiopathic PD cohort (P = 5.39 × 10-9 ), suggesting that every 5-year increase in DNAm-age acceleration is related to about a 6-year earlier onset. This link was replicated in an independent idiopathic PD cohort (P = 1.91 × 10-10 ). In each cohort, the faster-aging group has an increased hazard for an earlier onset (up to 255%). CONCLUSIONS: This study is the first to demonstrate that DNAm-age acceleration is related to PD age at onset, which could be considered in disease-modifying clinical trials. Future studies should evaluate the stability of DNAm-age acceleration over longer time periods, especially for phenoconverters from nonmanifesting to manifesting individuals. © 2022 International Parkinson and Movement Disorder Society.

α-Synuclein molecular behavior and nigral proteomic profiling distinguish subtypes of Lewy body disorders.

Lewy body disorders (LBD), characterized by the deposition of misfolded α-synuclein (α-Syn), are clinically heterogeneous. Although the distribution of α-Syn correlates with the predominant clinical features, the burden of pathology does not fully explain the observed variability in clinical presentation and rate of disease progression. We hypothesized that this heterogeneity might reflect α-Syn molecular diversity, between both patients and different brain regions. Using an ultra-sensitive assay, we evaluated α-Syn seeding in 8 brain regions from 30 LBD patients with different clinical phenotypes and disease durations. Comparing seeding across the clinical phenotypes revealed that hippocampal α-Syn from patients with a cognitive-predominant phenotype had significantly higher seeding capacity than that derived from patients with a motor-predominant phenotype, whose nigral-derived α-Syn in turn had higher seeding capacity than that from cognitive-predominant patients. Interestingly, α-Syn from patients with rapid disease progression (< 3 years to development of advanced disease) had the highest nigral seeding capacity of all the patients included. To validate these findings and explore factors underlying seeding heterogeneity, we performed in vitro toxicity assays, and detailed neuropathological and biochemical examinations. Furthermore, and for the first time, we performed a proteomic-wide profiling of the substantia nigra from 5 high seeder and 5 low seeder patients. The proteomic data suggests a significant disruption in mitochondrial function and lipid metabolism in high seeder cases compared to the low seeders. These observations suggest that distinct molecular populations of α-Syn may contribute to heterogeneity in phenotypes and progression rates in LBD and imply that effective therapeutic strategies might need to be directed at an ensemble of differently misfolded α-Syn species, with the relative contribution of their differing impacts accounting for heterogeneity in the neurodegenerative process.

Alpha-Synuclein Targeting Therapeutics for Parkinson's Disease and Related Synucleinopathies.

α-Synuclein (asyn) is a key pathogenetic factor in a group of neurodegenerative diseases generically known as synucleinopathies, including Parkinson's disease (PD), dementia with Lewy bodies (DLB) and multiple system atrophy (MSA). Although the initial triggers of pathology and progression are unclear, multiple lines of evidence support therapeutic targeting of asyn in order to limit its prion-like misfolding. Here, we review recent pre-clinical and clinical work that offers promising treatment strategies to sequester, degrade, or silence asyn expression as a means to reduce the levels of seed or substrate. These diverse approaches include removal of aggregated asyn with passive or active immunization or by expression of vectorized antibodies, modulating kinetics of misfolding with small molecule anti-aggregants, lowering asyn gene expression by antisense oligonucleotides or inhibitory RNA, and pharmacological activation of asyn degradation pathways. We also discuss recent technological advances in combining low intensity focused ultrasound with intravenous microbubbles to transiently increase blood-brain barrier permeability for improved brain delivery and target engagement of these large molecule anti-asyn biologics.

Patterns of Mixed Pathologies in Down Syndrome.

BACKGROUND: Down syndrome (DS) is frequently associated with Alzheimer's disease (AD)-related neuropathological changes. There are few observations on the spectrum of mixed proteinopathies in DS patients. OBJECTIVE: This study aimed to evaluate multiple disease-associated proteinopathies in a series of DS cases. METHODS: We analyzed the distribution of neurodegenerative disease associated proteins in postmortem brain samples from 11 DS cases (6 females, median age 57, range 38-66 years). Sections were stained for phosphorylated tau, 3-repeat and 4-repeat tau, amyloid-ß, alpha synuclein, phosphorylated TDP-43, and p62. A comprehensive anatomical mapping and staging were applied for all proteins. RESULTS: Tau and amyloid-ß pathology was prevalent in all cases and compatible with that typically seen in AD with some subtle deviations. Four of 11 cases presented with Lewy-related pathology (LRP). Two cases followed the Braak staging (stage 4 and 5) whereas 2 cases presented with an atypical distribution. Two cases showed limbic predominant age-related TDP-43 encephalopathy (LATE) (stage 1 and stage 2) neuropathologic change. Two cases exhibited aging-related tau astrogliopathy (ARTAG). CONCLUSION: In addition to subtle deviations from AD regarding the morphology of amyloid-ß deposition and distribution of neuronal tau pathology, we find that the spectrum of mixed-pathologies in DS show distinctive features such as deviations from the Braak staging of LRP and that LATE neuropathologic change and ARTAG pathology can be seen in individuals younger than in sporadic AD cases. Our observations support the notion that DS has distinctive pathogenic pathways from sporadic AD.

Alpha-synuclein seeding shows a wide heterogeneity in multiple system atrophy.

BACKGROUND: Multiple system atrophy (MSA) is a neurodegenerative condition characterized by variable combinations of parkinsonism, autonomic failure, cerebellar ataxia and pyramidal features. Although the distribution of synucleinopathy correlates with the predominant clinical features, the burden of pathology does not fully explain observed differences in clinical presentation and rate of disease progression. We hypothesized that the clinical heterogeneity in MSA is a consequence of variability in the seeding activity of α-synuclein both between different patients and between different brain regions. METHODS: The reliable detection of α-synuclein seeding activity derived from MSA using cell-free amplification assays remains challenging. Therefore, we conducted a systematic evaluation of 168 different reaction buffers, using an array of pH and salts, seeded with fully characterized brain homogenates from one MSA and one PD patient. We then validated the two conditions that conferred the optimal ability to discriminate between PD- and MSA-derived samples in a larger cohort of 40 neuropathologically confirmed cases, including 15 MSA. Finally, in a subset of brains, we conducted the first multi-region analysis of seeding behaviour in MSA. RESULTS: Using our novel buffer conditions, we show that the physicochemical factors that govern the in vitro amplification of α-synuclein can be tailored to generate strain-specific reaction buffers that can be used to reliably study the seeding capacity from MSA-derived α-synuclein. Using this novel approach, we were able to sub-categorize the 15 MSA brains into 3 groups: high, intermediate and low seeders. To further demonstrate heterogeneity in α-synuclein seeding in MSA, we conducted a comprehensive multi-regional evaluation of α-synuclein seeding in 13 different regions from 2 high seeders, 2 intermediate seeders and 2 low seeders. CONCLUSIONS: We have identified unexpected differences in seed-competent α-synuclein across a cohort of neuropathologically comparable MSA brains. Furthermore, our work has revealed a substantial heterogeneity in seeding activity, driven by the PBS-soluble α-synuclein, between different brain regions of a given individual that goes beyond immunohistochemical observations. Our observations pave the way for future subclassification of MSA, which exceeds conventional clinical and neuropathological phenotyping and considers the structural and biochemical heterogeneity of α-synuclein present. Finally, our methods provide an experimental framework for the development of vitally needed, rapid and sensitive diagnostic assays for MSA.

Axial Impairment Following Deep Brain Stimulation in Parkinson's Disease: A Surgicogenomic Approach.

BACKGROUND: Postoperative outcome following deep brain stimulation (DBS) of the subthalamic nucleus is variable, particularly with respect to axial motor improvement. We hypothesized a genetic underpinning to the response to surgical intervention, termed "surgicogenomics". OBJECTIVE: We aimed to identify genetic variants associated with clinical heterogeneity in DBS outcome of Parkinson's disease (PD) patients that could then be applied clinically to target selection leading to improved surgical outcome. METHODS: Retrospective clinical data was extracted from 150 patient's charts. Each individual was genotyped using the genome-wide NeuroX array tailored to study neurologic diseases. Genetic data were clustered based on surgical outcome assessed by comparing pre- and post-operative scores of levodopa equivalent daily dose and axial impairment at one and five years post-surgery. Allele frequencies were compared between patients with excellent vs. moderate/poor outcomes grouped using a priori defined cut-offs. We analyzed common variants, burden of rare coding variants, and PD polygenic risk score. RESULTS: NeuroX identified 2,917 polymorphic markers at 113 genes mapped to known PD loci. The gene-burden analyses of 202 rare nonsynonymous variants suggested a nominal association of axial impairment with 14 genes (most consistent with CRHR1, IP6K2, and PRSS3). The strongest association with surgical outcome was detected between a reduction in levodopa equivalent daily dose and common variations tagging two linkage disequilibrium blocks with SH3GL2. CONCLUSION: Once validated in independent populations, our findings may be implemented to improve patient selection for DBS in PD.

Small molecule inhibitors of α-synuclein oligomers identified by targeting early dopamine-mediated motor impairment in C. elegans.

BACKGROUND: Parkinson's disease is a disabling neurodegenerative movement disorder characterized by dopaminergic neuron loss induced by α-synuclein oligomers. There is an urgent need for disease-modifying therapies for Parkinson's disease, but drug discovery is challenged by lack of in vivo models that recapitulate early stages of neurodegeneration. Invertebrate organisms, such as the nematode worm Caenorhabditis elegans, provide in vivo models of human disease processes that can be instrumental for initial pharmacological studies. METHODS: To identify early motor impairment of animals expressing α-synuclein in dopaminergic neurons, we first used a custom-built tracking microscope that captures locomotion of single C. elegans with high spatial and temporal resolution. Next, we devised a method for semi-automated and blinded quantification of motor impairment for a population of simultaneously recorded animals with multi-worm tracking and custom image processing. We then used genetic and pharmacological methods to define the features of early motor dysfunction of α-synuclein-expressing C. elegans. Finally, we applied the C. elegans model to a drug repurposing screen by combining it with an artificial intelligence platform and cell culture system to identify small molecules that inhibit α-synuclein oligomers. Screen hits were validated using in vitro and in vivo mammalian models. RESULTS: We found a previously undescribed motor phenotype in transgenic α-synuclein C. elegans that correlates with mutant or wild-type α-synuclein protein levels and results from dopaminergic neuron dysfunction, but precedes neuronal loss. Together with artificial intelligence-driven in silico and in vitro screening, this C. elegans model identified five compounds that reduced motor dysfunction induced by α-synuclein. Three of these compounds also decreased α-synuclein oligomers in mammalian neurons, including rifabutin which has not been previously investigated for Parkinson's disease. We found that treatment with rifabutin reduced nigrostriatal dopaminergic neurodegeneration due to α-synuclein in a rat model. CONCLUSIONS: We identified a C. elegans locomotor abnormality due to dopaminergic neuron dysfunction that models early α-synuclein-mediated neurodegeneration. Our innovative approach applying this in vivo model to a multi-step drug repurposing screen, with artificial intelligence-driven in silico and in vitro methods, resulted in the discovery of at least one drug that may be repurposed as a disease-modifying therapy for Parkinson's disease.

Exposure to Phosphoglycerate Kinase 1 Activators and Incidence of Parkinson's Disease.

BACKGROUND: Preclinical and epidemiological data suggest that phosphoglycerate kinase 1 activators could have neuroprotective properties and prevent PD. OBJECTIVES: The objective of this study was to compare the association between increased use of phosphoglycerate kinase 1 activators and increased use of tamsulosin with PD incidence. METHODS: Our retrospective cohort study included men older than age 66 years newly exposed to phosphoglycerate kinase 1 activators or tamsulosin and compared their PD incidence, using health care administrative data of Ontario, Canada. RESULTS: Among 265,745 men, each additional year of cumulative use of phosphoglycerate kinase 1 activators or tamsulosin was associated with 6% and 8% reduction, respectively, in the hazard of PD incidence. These hazards were not significantly different (P = 0.2094). A secondary analysis with the observation window starting after 6 months and 1 and 2 years showed similar results. CONCLUSIONS: Increasing exposure to phosphoglycerate kinase 1 activators and tamsulosin were both associated with small reductions in PD incidence. These results support further investigation of phosphoglycerate kinase 1 activators and tamsulosin for possible PD disease-modifying properties. © 2021 International Parkinson and Movement Disorder Society.

Short-term deceleration capacity of heart rate: a sensitive marker of cardiac autonomic dysfunction in idiopathic Parkinson's disease.

PURPOSE: Cardiac autonomic dysfunction in idiopathic Parkinson's disease (PD) manifests as reduced heart rate variability (HRV). In the present study, we explored the deceleration capacity of heart rate (DC) in patients with idiopathic PD, an advanced HRV marker that has proven clinical utility. METHODS: Standard and advanced HRV measures derived from 7-min electrocardiograms in 20 idiopathic PD patients and 27 healthy controls were analyzed. HRV measures were compared using regression analysis, controlling for age, sex, and mean heart rate. RESULTS: Significantly reduced HRV was found only in the subcohort of PD patients older than 60 years. Low- frequency power and global HRV measures were lower in patients than in controls, but standard beat-to-beat HRV markers (i.e., rMSSD and high-frequency power) were not significantly different between groups. DC was significantly reduced in the subcohort of PD patients older than 60 years compared to controls. CONCLUSIONS: Deceleration-related oscillations of HRV were significantly reduced in the older PD patients compared to healthy controls, suggesting that short-term DC may be a sensitive marker of cardiac autonomic dysfunction in PD. DC may be complementary to traditional markers of short-term HRV for the evaluation of autonomic modulation in PD. Further study to examine the association between DC and cardiac adverse events in PD is needed to clarify the clinical relevance of DC in this population.

Using artificial intelligence to identify anti-hypertensives as possible disease modifying agents in Parkinson's disease.

PURPOSE: Drug repurposing is an effective means of increasing treatment options for diseases, however identifying candidate molecules for the indication of interest from the thousands of approved drugs is challenging. We have performed a computational analysis of published literature to rank existing drugs according to predicted ability to reduce alpha synuclein (aSyn) oligomerization and analyzed real-world data to investigate the association between exposure to highly ranked drugs and PD. METHODS: Using IBM Watson for Drug Discoveryâ (WDD) we identified several antihypertensive drugs that may reduce aSyn oligomerization. Using IBM MarketScanâ Research Databases we constructed a cohort of individuals with incident hypertension. We conducted univariate and multivariate Cox proportional hazard analyses (HR) with exposure as a time-dependent covariate. Diuretics were used as the referent group. Age at hypertension diagnosis, sex, and several comorbidities were included in multivariate analyses. RESULTS: Multivariate results revealed inverse associations for time to PD diagnosis with exposure to the combination of the combination of angiotensin receptor II blockers (ARBs) and dihydropyridine calcium channel blockers (DHP-CCB) (HR = 0.55, p < 0.01) and angiotensin converting enzyme inhibitors (ACEi) and diuretics (HR = 0.60, p-value <0.01). Increased risk was observed with exposure to alpha-blockers alone (HR = 1.81, p < 0.001) and the combination of alpha-blockers and CCB (HR = 3.17, p < 0.05). CONCLUSIONS: We present evidence that a computational approach can efficiently identify leads for disease-modifying drugs. We have identified the combination of ARBs and DHP-CCBs as of particular interest in PD.