Alpha-Synuclein Gene Alterations Modulate Tyrosine Hydroxylase in Human iPSC-Derived Neurons in a Parkinson's Disease Animal Model.

Parkinson's disease (PD) caused by SNCA gene triplication (3XSNCA) leads to early onset, rapid progression, and often dementia. Understanding the impact of 3XSNCA and its absence is crucial. This study investigates the differentiation of human induced pluripotent stem cell (hiPSC)-derived floor-plate progenitors into dopaminergic neurons. Three different genotypes were evaluated in this study: patient-derived hiPSCs with 3XSNCA, a gene-edited isogenic line with a frame-shift mutation on all SNCA alleles (SNCA 4KO), and a normal wild-type control. Our aim was to assess how the substantia nigra pars compacta (SNpc) microenvironment, damaged by 6-hydroxydopamine (6-OHDA), influences tyrosine hydroxylase-positive (Th+) neuron differentiation in these genetic variations. This study confirms successful in vitro differentiation into neuronal lineage in all cell lines. However, the SNCA 4KO line showed unusual LIM homeobox transcription factor 1 alpha (Lmx1a) extranuclear distribution. Crucially, both 3XSNCA and SNCA 4KO lines had reduced Th+ neuron expression, despite initial successful neuronal differentiation after two months post-transplantation. This indicates that while the SNpc environment supports early neuronal survival, SNCA gene alterations-either amplification or knock-out-negatively impact Th+ dopaminergic neuron maturation. These findings highlight SNCA's critical role in PD and underscore the value of hiPSC models in studying neurodegenerative diseases.

Identification and validation of N-acetylputrescine in combination with non-canonical clinical features as a Parkinson's disease biomarker panel.

Parkinson's disease is a progressive neurodegenerative disorder in which loss of dopaminergic neurons in the substantia nigra results in a clinically heterogeneous group with variable motor and non-motor symptoms with a degree of misdiagnosis. Only 3-25% of sporadic Parkinson's patients present with genetic abnormalities that could represent a risk factor, thus environmental, metabolic, and other unknown causes contribute to the pathogenesis of Parkinson's disease, which highlights the critical need for biomarkers. In the present study, we prospectively collected and analyzed plasma samples from 194 Parkinson's disease patients and 197 age-matched non-diseased controls. N-acetyl putrescine (NAP) in combination with sense of smell (B-SIT), depression/anxiety (HADS), and acting out dreams (RBD1Q) clinical measurements demonstrated combined diagnostic utility. NAP was increased by 28% in Parkinsons disease patients and exhibited an AUC of 0.72 as well as an OR of 4.79. The clinical and NAP panel demonstrated an area under the curve, AUC = 0.9 and an OR of 20.4. The assessed diagnostic panel demonstrates combinatorial utility in diagnosing Parkinson's disease, allowing for an integrated interpretation of disease pathophysiology and highlighting the use of multi-tiered panels in neurological disease diagnosis.

Inactive S. aureus Cas9 downregulates alpha-synuclein and reduces mtDNA damage and oxidative stress levels in human stem cell model of Parkinson's disease.

Parkinson's disease (PD) is one of the most common neurodegenerative diseases, but no disease modifying therapies have been successful in clinical translation presenting a major unmet medical need. A promising target is alpha-synuclein or its aggregated form, which accumulates in the brain of PD patients as Lewy bodies. While it is not entirely clear which alpha-synuclein protein species is disease relevant, mere overexpression of alpha-synuclein in hereditary forms leads to neurodegeneration. To specifically address gene regulation of alpha-synuclein, we developed a CRISPR interference (CRISPRi) system based on the nuclease dead S. aureus Cas9 (SadCas9) fused with the transcriptional repressor domain Krueppel-associated box to controllably repress alpha-synuclein expression at the transcriptional level. We screened single guide (sg)RNAs across the SNCA promoter and identified several sgRNAs that mediate downregulation of alpha-synuclein at varying levels. CRISPRi downregulation of alpha-synuclein in iPSC-derived neuronal cultures from a patient with an SNCA genomic triplication showed functional recovery by reduction of oxidative stress and mitochondrial DNA damage. Our results are proof-of-concept in vitro for precision medicine by targeting the SNCA gene promoter. The SNCA CRISPRi approach presents a new model to understand safe levels of alpha-synuclein downregulation and a novel therapeutic strategy for PD and related alpha-synucleinopathies.

Automated 384-well SYBR Green Expression Array for Optimization of Human Induced Pluripotent Stem Cell Differentiation.

Cell populations and tissues exhibit unique gene expression profiles, which allow for characterizing and distinguishing cellular subtypes. Monitoring gene expression of cell type-specific markers can indicate cell status such as proliferation, stress, quiescence, or maturation. Quantitative reverse transcriptase PCR (qRT-PCR) allows quantifying RNA expression of cell type-specific markers and distinguishing one cell type from another. However, qRT-PCR methods such as TaqMan technology require fluorescent reporters to characterize target genes and are challenging to scale up as they need different probes for each reaction. Bulk or single-cell RNA transcriptomics is time-consuming and expensive. Processing RNA sequencing data can take several weeks, which is not optimal for quality control and monitoring gene expression, e.g., during a differentiation paradigm of induced pluripotent stem cells (iPSCs) into a specialized cell type. A more cost-effective assay is based on SYBR Green technology. SYBR Green is a nucleic acid dye that binds to double-stranded DNA, absorbs blue light at 497 nm, and emits green light at 520 nm up to 1,000-fold upon intercalation with double-stranded DNA. Amplification of a region of interest can be quantified based on the level of fluorescence intensity when normalized to a housekeeping gene and compared to control conditions. Previously, we established a SYBR Green qRT-PCR protocol to characterize samples using a limited set of markers plated on a 96-well plate. Here, we optimize the process and increase throughput to a 384-well format and compare mRNA expression to distinguish iPSC-derived neuronal subtypes from each other by increasing the number of genes, cell types, and differentiation time points. In this protocol, we develop the following: i) using the command-line version of Primer3 software, we design primers more easily and quickly for the gene of interest; ii) using a 384-well plate format, electronic multichannel pipettes, and pipetting robots, we analyze four times more genes on a single plate while using the same volume of reagents as in a 96-well plate. The advantages of this protocol are the increased throughput of this SYBR Green assay while limiting pipetting errors/inconsistencies, reagent use, cost, and time. Graphical overview.

Nuclease-dead S. aureus Cas9 downregulates alpha-synuclein and reduces mtDNA damage and oxidative stress levels in patient-derived stem cell model of Parkinson's disease.

Parkinson's disease (PD) is one of the most common neurodegenerative diseases, but no disease modifying therapies have been successful in clinical translation presenting a major unmet medical need. A promising target is alpha-synuclein or its aggregated form, which accumulates in the brain of PD patients as Lewy bodies. While it is not entirely clear which alpha-synuclein protein species is disease relevant, mere overexpression of alpha-synuclein in hereditary forms leads to neurodegeneration. To specifically address gene regulation of alpha-synuclein, we developed a CRISPR interference (CRISPRi) system based on the nuclease dead S. aureus Cas9 (SadCas9) fused with the transcriptional repressor domain Krueppel-associated box to controllably repress alpha-synuclein expression at the transcriptional level. We screened single guide (sg)RNAs across the SNCA promoter and identified several sgRNAs that mediate downregulation of alpha-synuclein at varying levels. CRISPRi downregulation of alpha-synuclein in iPSC-derived neuronal cultures from a patient with an SNCA genomic triplication showed functional recovery by reduction of oxidative stress and mitochondrial DNA damage. Our results are proof-of-concept in vitro for precision medicine by targeting the SNCA gene promoter. The SNCA CRISPRi approach presents a new model to understand safe levels of alpha-synuclein downregulation and a novel therapeutic strategy for PD and related alpha-synucleinopathies.

Embryoid Body Cells from Human Embryonic Stem Cells Overexpressing Dopaminergic Transcription Factors Survive and Initiate Neurogenesis via Neural Rosettes in the Substantia Nigra.

Transplantation of immature dopaminergic neurons or neural precursors derived from embryonic stem cells (ESCs) into the substantia nigra pars compacta (SNpc) is a potential therapeutic approach for functional restitution of the nigrostriatal pathway in Parkinson's disease (PD). However, further studies are needed to understand the effects of the local microenvironment on the transplanted cells to improve survival and specific differentiation in situ. We have previously reported that the adult SNpc sustains a neurogenic microenvironment. Non-neuralized embryoid body cells (EBCs) from mouse ESCs (mESCs) overexpressing the dopaminergic transcription factor Lmx1a gave rise to many tyrosine hydroxylase (Th+) cells in the intact and damaged adult SNpc, although only for a short-term period. Here, we extended our study by transplanting EBCs from genetically engineered naive human ESC (hESC), overexpressing the dopaminergic transcription factors LMX1A, FOXA2, and OTX2 (hESC-LFO), in the SNpc. Unexpectedly, no graft survival was observed in wild-type hESC EBCs transplants, whereas hESC-LFO EBCs showed viability in the SNpc. Interestingly, neural rosettes, a developmental hallmark of neuroepithelial tissue, emerged at 7- and 15-days post-transplantation (dpt) from the hESC-LFO EBCs. Neural rosettes expressed specification dopaminergic markers (Lmx1a, Otx2), which gave rise to several Th+ cells at 30 dpt. Our results suggest that the SNpc enables the robust initiation of neural differentiation of transplanted human EBCs prompted to differentiate toward the midbrain dopaminergic phenotype.

Early embryonic lethality in complex I associated p.L104P Nubpl mutant mice.

BACKGROUND: Variants in the mitochondrial complex I assembly factor, NUBPL are associated with a rare cause of complex I deficiency mitochondrial disease. Patients affected by complex I deficiency harboring homozygous NUBPL variants typically have neurological problems including seizures, intellectual disability, and ataxia associated with cerebellar hypoplasia. Thus far only 19 cases have been reported worldwide, and no treatment is available for this rare disease. METHODS: To investigate the pathogenesis of NUBPL-associated complex I deficiency, and for translational studies, we generated a knock-in mouse harboring a patient-specific variant Nubpl c.311T>C; p. L104P reported in three families. RESULTS: Similar to Nubpl global knockout mice, the Nubpl p. L104P homozygous mice are lethal at embryonic day E10.5, suggesting that the Nubpl p. L104P variant is likely a hypomorph allele. Given the recent link between Parkinson's disease and loss-of-function NUBPL variants, we also explored aging-related behaviors and immunocytochemical changes in Nubpl hemizygous mice and did not find significant behavioral and pathological changes for alpha-synuclein and oxidative stress markers . CONCLUSION: Our data suggest that homozygotes with Nubpl variants, similar to the null mice, are lethal, and heterozygotes are phenotypically and neuropathologically normal. We propose that a tissue-specific knockout strategy is required to establish a mouse model of Nubpl-associated complex I deficiency disorder for future mechanistic and translational studies.

ATTCT and ATTCC repeat expansions in the ATXN10 gene affect disease penetrance of spinocerebellar ataxia type 10.

Spinocerebellar ataxia type 10 (SCA10) is an autosomal-dominant disorder caused by an expanded pentanucleotide repeat in the ATXN10 gene. This repeat expansion, when fully penetrant, has a size of 850-4,500 repeats. It has been shown that the repeat composition can be a modifier of disease, e.g., seizures. Here, we describe a Mexican kindred in which we identified both pure (ATTCT)n and mixed (ATTCT)n-(ATTCC)n expansions in the same family. We used amplification-free targeted sequencing and optical genome mapping to decipher the composition of these repeat expansions. We found a considerable degree of mosaicism of the repeat expansion. This mosaicism was confirmed in skin fibroblasts from individuals with ATXN10 expansions with RNAScope in situ hybridization. All affected family members with the mixed ATXN10 repeat expansion showed typical clinical signs of spinocerebellar ataxia and epilepsy. In contrast, individuals with the pure ATXN10 expansion present with Parkinson's disease or are unaffected, even in individuals more than 20 years older than the average age at onset for SCA10. Our findings suggest that the pure (ATTCT)n expansion is non-pathogenic, while repeat interruptions, e.g., (ATTCC)n, are necessary to cause SCA10. This mechanism has been recently described for several other repeat expansions including SCA31 (BEAN1), SCA37 (DAB1), and three loci for benign adult familial myoclonic epilepsy BAFME (SAMD12, TNRC6A, RAPGEF2). Therefore, long-read sequencing and optical genome mapping of the entire genomic structure of repeat expansions are critical for clinical practice and genetic counseling, as variations in the repeat can affect disease penetrance, symptoms, and disease trajectory.

Gene Expression Analysis in Stem Cell-derived Cortical Neuronal Cultures Using Multi-well SYBR Green Quantitative PCR Arrays.

To optimize differentiation protocols for stem cell-based in vitro modeling applications, it is essential to assess the change in gene expression during the differentiation process. This allows controlling its differentiation efficiency into the target cell types. While RNA transcriptomics provides detail at a larger scale, timing and cost are prohibitive to include such analyses in the optimization process. In contrast, expression analysis of individual genes is cumbersome and lengthy. Here, we developed a versatile and cost-efficient SYBR Green array of 27 markers along with two housekeeping genes to quickly screen for differentiation efficiency of human induced pluripotent stem cells (iPSCs) into excitatory cortical neurons. We first identified relevant pluripotency, neuroprogenitor, and neuronal markers for the array by literature search, and designed primers with a product size of 80-120 bp length, an annealing temperature of 60°C, and minimal predicted secondary structures. We spotted combined forward and reverse primers on 96-well plates and dried them out overnight. These plates can be prepared in advance in batches and stored at room temperature until use. Next, we added the SYBR Green master mix and complementary DNA (cDNA) to the plate in triplicates, ran quantitative PCR (qPCR) on a Quantstudio 6 Flex, and analyzed results with QuantStudio software. We compared the expression of genes for pluripotency, neuroprogenitor cells, cortical neurons, and synaptic markers in a 96-well format at four different time points during the cortical differentiation. We found a sharp reduction of pluripotency genes within the first three days of pre-differentiation and a steady increase of neuronal markers and synaptic markers over time. In summary, we built a gene expression array that is customizable, fast, medium-throughput, and cost-efficient, ideally suited for optimization of differentiation protocols for stem cell-based in vitro modeling.

Multiplex imaging of human induced pluripotent stem cell-derived neurons with CO-Detection by indEXing (CODEX) technology.

BACKGROUND: Human induced pluripotent stem cell (iPSC) models have been hailed as a breakthrough for understanding disease and developing new therapeutics. The major advantage of iPSC-derived neurons is that they carry the genetic background of the donor, and as such could be more predictive for clinical translation. However, the development of these cell models is time-consuming and expensive and it is thus critical to maximize readout of markers for immunocytochemistry. One option is to use a highly multiplexed fluorescence imaging assay, like CO-Detection by indEXing (CODEX), which allows detection of 50 + targets in situ. NEW METHOD: This paper describes the development of CODEX in neuronal cell cultures derived from human iPSCs. RESULTS: We differentiated human iPSCs into mixed neuronal and glial cultures on glass coverslips. We then developed and optimized a panel of 21 antibodies to phenotype iPSC-derived neuronal subtypes of cortical, dopaminergic, and striatal neurons, as well as astrocytes, and pre-and postsynaptic proteins. COMPARISON WITH EXISTING METHODS: Compared to standard immunocytochemistry, CODEX oligo-conjugated fluorophores circumvent antibody host interactions and allow for highly customized multiplexing. CONCLUSION: We show that CODEX can be applied to iPSC neuronal cultures and developed fixation and staining protocols for the neurons to sustain the multiple wash-stain cycles of the technology. Furthermore, we demonstrate both cellular and subcellular resolution imaging of multiplexed markers in the same sample.

Isogenic human SNCA gene dosage induced pluripotent stem cells to model Parkinson's disease.

Alpha-synuclein overexpression and aggregation are critical factors in the pathogenesis of Parkinson's disease (PD). Clinical cases with alpha-synuclein (SNCA) multiplications or deletions indicate that gene expression levels are essential for neurodegeneration and neurodevelopment. Here, we developed an isogenic SNCA gene dosage model using CRISPR/Cas9 gene editing to introduce frameshift mutations into exon 2 of the SNCA coding region in human induced pluripotent stem cells (iPSCs) from a patient with an SNCA triplication. We derived and characterized clones with different frameshift mutations. This isogenic SNCA gene dosage panel will address the physiological and detrimental effects of varying alpha-synuclein expression levels.

Effect of LRRK2 protein and activity on stimulated cytokines in human monocytes and macrophages.

Leucine-rich-repeat kinase 2 (LRRK2), a potential therapeutic target for the treatment of Parkinson's disease (PD), is highly expressed in monocytes and macrophages and may play a role in the regulation of inflammatory pathways. To determine how LRRK2 protein levels and/or its activity modulate inflammatory cytokine/chemokine levels in human immune cells, isogenic human induced pluripotent stem cells (iPSC) with the LRRK2-activating G2019S mutation, wild-type LRRK2, and iPSC deficient in LRRK2 were differentiated to monocytes and macrophages and stimulated with inflammatory toll-like receptor (TLR) agonists in the presence and absence of LRRK2 kinase inhibitors. The effect of LRRK2 inhibitors and the effect of increasing LRRK2 levels with interferon gamma on TLR-stimulated cytokines were also assessed in primary peripheral blood-derived monocytes. Monocytes and macrophages with the LRRK2 G2019S mutation had significantly higher levels of cytokines and chemokines in tissue culture media following stimulation with TLR agonists compared to isogenic controls. Knockout of LRRK2 impaired phagocytosis but did not significantly affect TLR-mediated cytokine levels. Interferon gamma significantly increased the levels of LRRK2 and phosphorylation of its downstream Rab10 substrate, and potentiated TLR-mediated cytokine levels. LRRK2 kinase inhibitors did not have a major effect on TLR-stimulated cytokine levels. Results suggest that the LRRK2 G2019S mutation may potentiate inflammation following activation of TLRs. However, this was not dependent on LRRK2 kinase activity. Indeed, LRRK2 kinase inhibitors had little effect on TLR-mediated inflammation under the conditions employed in this study.

Phenotypic Heterogeneity among GBA p.R202X Carriers in Lewy Body Spectrum Disorders.

We describe the clinical and neuropathologic features of patients with Lewy body spectrum disorder (LBSD) carrying a nonsense variant, c.604C>T; p.R202X, in the glucocerebrosidase 1 (GBA) gene. While this GBA variant is causative for Gaucher's disease, the pathogenic role of this mutation in LBSD is unclear. Detailed neuropathologic evaluation was performed for one index case and a structured literature review of other GBA p.R202X carriers was conducted. Through the systematic literature search, we identified three additional reported subjects carrying the same GBA mutation, including one Parkinson's disease (PD) patient with early disease onset, one case with neuropathologically-verified LBSD, and one unaffected relative of a Gaucher's disease patient. Among the affected subjects carrying the GBA p.R202X, all males were diagnosed with Lewy body dementia, while the two females presented as PD. The clinical penetrance of GBA p.R202X in LBSD patients and families argues strongly for a pathogenic role for this variant, although presenting with a striking phenotypic heterogeneity of clinical and pathological features.

Embryoid Body Formation from Mouse and Human Pluripotent Stem Cells for Transplantation to Study Brain Microenvironment and Cellular Differentiation.

Human embryonic stem cell (hESC) and human-induced pluripotent stem cell (hiPSC) technologies have a critical role in regenerative strategies for personalized medicine. Both share the ability to differentiate into almost any cell type of the human body. The study of their properties and clinical applications requires the development of robust and reproducible cell culture paradigms that direct cell differentiation toward a specific phenotype in vitro and in vivo. Our group evaluated the potential of mouse ESCs (mESCs), hESCs, and hiPSCs (collectively named pluripotent stem cells, PSCs) to analyze brain microenvironments through the use of embryoid body (EB)-derived cells from these cell sources. EB are cell aggregates in 3D culture conditions that recapitulate embryonic development. Our approach focuses on studying the midbrain dopaminergic phenotype and transplanting EB into the substantia nigra pars compacta (SNpc) in a Parkinson's disease rodent model. Here, we describe cell culture protocols for EB generation from PSCs that show significant in vivo differentiation toward dopaminergic neurons.

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.

Genomewide Association Studies of LRRK2 Modifiers of Parkinson's Disease.

OBJECTIVE: The aim of this study was to search for genes/variants that modify the effect of LRRK2 mutations in terms of penetrance and age-at-onset of Parkinson's disease. METHODS: We performed the first genomewide association study of penetrance and age-at-onset of Parkinson's disease in LRRK2 mutation carriers (776 cases and 1,103 non-cases at their last evaluation). Cox proportional hazard models and linear mixed models were used to identify modifiers of penetrance and age-at-onset of LRRK2 mutations, respectively. We also investigated whether a polygenic risk score derived from a published genomewide association study of Parkinson's disease was able to explain variability in penetrance and age-at-onset in LRRK2 mutation carriers. RESULTS: A variant located in the intronic region of CORO1C on chromosome 12 (rs77395454; p value = 2.5E-08, beta = 1.27, SE = 0.23, risk allele: C) met genomewide significance for the penetrance model. Co-immunoprecipitation analyses of LRRK2 and CORO1C supported an interaction between these 2 proteins. A region on chromosome 3, within a previously reported linkage peak for Parkinson's disease susceptibility, showed suggestive associations in both models (penetrance top variant: p value = 1.1E-07; age-at-onset top variant: p value = 9.3E-07). A polygenic risk score derived from publicly available Parkinson's disease summary statistics was a significant predictor of penetrance, but not of age-at-onset. INTERPRETATION: This study suggests that variants within or near CORO1C may modify the penetrance of LRRK2 mutations. In addition, common Parkinson's disease associated variants collectively increase the penetrance of LRRK2 mutations. ANN NEUROL 2021;90:82-94.

Genetic and Environmental Factors Influence the Pleomorphy of LRRK2 Parkinsonism.

Missense mutations in the LRRK2 gene were first identified as a pathogenic cause of Parkinson's disease (PD) in 2004. Soon thereafter, a founder mutation in LRRK2, p.G2019S (rs34637584), was described, and it is now estimated that there are approximately 100,000 people worldwide carrying this risk variant. While the clinical presentation of LRRK2 parkinsonism has been largely indistinguishable from sporadic PD, disease penetrance and age at onset can be quite variable. In addition, its neuropathological features span a wide range from nigrostriatal loss with Lewy body pathology, lack thereof, or atypical neuropathology, including a large proportion of cases with concomitant Alzheimer's pathology, hailing LRRK2 parkinsonism as the "Rosetta stone" of parkinsonian disorders, which provides clues to an understanding of the different neuropathological trajectories. These differences may result from interactions between the LRRK2 mutant protein and other proteins or environmental factors that modify LRRK2 function and, thereby, influence pathobiology. This review explores how potential genetic and biochemical modifiers of LRRK2 function may contribute to the onset and clinical presentation of LRRK2 parkinsonism. We review which genetic modifiers of LRRK2 influence clinical symptoms, age at onset, and penetrance, what LRRK2 mutations are associated with pleomorphic LRRK2 neuropathology, and which environmental modifiers can augment LRRK2 mutant pathophysiology. Understanding how LRRK2 function is influenced and modulated by other interactors and environmental factors-either increasing toxicity or providing resilience-will inform targeted therapeutic development in the years to come. This will allow the development of disease-modifying therapies for PD- and LRRK2-related neurodegeneration.

Loss-of-Function NUBPL Mutation May Link Parkinson's Disease to Recessive Complex I Deficiency.

In an unbiased genome-wide screen for copy number variants (CNVs) on a cohort of Parkinson's disease (PD) patients, we identified in one patient a complex chromosomal rearrangement involving the nucleotide binding protein-like (NUBPL) gene on chromosome 14q12. We noted that mutations in the NUBPL gene had been reported as causing autosomal recessive (AR) mitochondrial Complex I (CI) deficiency in children. The precise breakpoints of the rearrangement in our PD case were found to be identical to those described in a patient with AR CI deficiency who also harbored a second pathogenic mutation in NUBPL. Mitochondrial dysfunction has long been considered a strong contributor to PD, and there is substantial evidence that decreased CI activity plays a central role in PD pathogenesis. We hypothesize that pathogenic NUBPL variants may increase the risk for PD analogous to variants in the glucosylceramidase beta (GBA) gene that increase the risk of developing PD in heterozygous carriers.

The Role of Alpha-Synuclein and Other Parkinson's Genes in Neurodevelopmental and Neurodegenerative Disorders.

Neurodevelopmental and late-onset neurodegenerative disorders present as separate entities that are clinically and neuropathologically quite distinct. However, recent evidence has highlighted surprising commonalities and converging features at the clinical, genomic, and molecular level between these two disease spectra. This is particularly striking in the context of autism spectrum disorder (ASD) and Parkinson's disease (PD). Genetic causes and risk factors play a central role in disease pathophysiology and enable the identification of overlapping mechanisms and pathways. Here, we focus on clinico-genetic studies of causal variants and overlapping clinical and cellular features of ASD and PD. Several genes and genomic regions were selected for our review, including SNCA (alpha-synuclein), PARK2 (parkin RBR E3 ubiquitin protein ligase), chromosome 22q11 deletion/DiGeorge region, and FMR1 (fragile X mental retardation 1) repeat expansion, which influence the development of both ASD and PD, with converging features related to synaptic function and neurogenesis. Both PD and ASD display alterations and impairments at the synaptic level, representing early and key disease phenotypes, which support the hypothesis of converging mechanisms between the two types of diseases. Therefore, understanding the underlying molecular mechanisms might inform on common targets and therapeutic approaches. We propose to re-conceptualize how we understand these disorders and provide a new angle into disease targets and mechanisms linking neurodevelopmental disorders and neurodegeneration.

Nonsteroidal Anti-inflammatory Use and LRRK2 Parkinson's Disease Penetrance.

BACKGROUND: The penetrance of leucine rich repeat kinase 2 (LRRK2) mutations is incomplete and may be influenced by environmental and/or other genetic factors. Nonsteroidal anti-inflammatory drugs (NSAIDs) are known to reduce inflammation and may lower Parkinson's disease (PD) risk, but their role in LRRK2-associated PD is unknown. OBJECTIVES: The objective of this study is to evaluate the association of regular NSAID use and LRRK2-associated PD. METHODS: Symptomatic ("LRRK2-PD") and asymptomatic ("LRRK2-non-PD") participants with LRRK2 G2019S, R1441X, or I2020T variants (definitely pathogenic variant carriers) or G2385R or R1628P variants (risk variant carriers) from 2 international cohorts provided information on regular ibuprofen and/or aspirin use (≥2 pills/week for ≥6 months) prior to the index date (diagnosis date for PD, interview date for non-PD). Multivariate logistic regression was used to evaluate the relationship between regular NSAID use and PD for any NSAID, separately for ibuprofen and aspirin in all carriers and separately in pathogenic and risk variant groups. RESULTS: A total of 259 LRRK2-PD and 318 LRRK2-non-PD participants were enrolled. Regular NSAID use was associated with reduced odds of PD in the overall cohort (odds ratio [OR], 0.34; 95% confidence interval [CI], 0.21-0.57) and in both pathogenic and risk variant carriers (ORPathogenic , 0.38; 95% CI, 0.21-0.67 and ORRiskVariant , 0.19; 95% CI, 0.04-0.99). Similar associations were observed for ibuprofen and aspirin separately (ORIbuprofen , 0.19; 95% CI, 0.07-0.50 and ORAspirin , 0.51; 95% CI, 0.28-0.91). CONCLUSIONS: Regular NSAID use may be associated with reduced penetrance in LRRK2-associated PD. The LRRK2 protein is involved in inflammatory pathways and appears to be modulated by regular anti-inflammatory use. Longitudinal observational and interventional studies of NSAID exposure and LRRK2-PD are needed to confirm this association. © 2020 International Parkinson and Movement Disorder Society.