Advancements in APOE and dementia research: Highlights from the 2023 AAIC Advancements: APOE conference.

INTRODUCTION: The apolipoprotein E gene (APOE) is an established central player in the pathogenesis of Alzheimer's disease (AD), with distinct apoE isoforms exerting diverse effects. apoE influences not only amyloid-beta and tau pathologies but also lipid and energy metabolism, neuroinflammation, cerebral vascular health, and sex-dependent disease manifestations. Furthermore, ancestral background may significantly impact the link between APOE and AD, underscoring the need for more inclusive research. METHODS: In 2023, the Alzheimer's Association convened multidisciplinary researchers at the "AAIC Advancements: APOE" conference to discuss various topics, including apoE isoforms and their roles in AD pathogenesis, progress in apoE-targeted therapeutic strategies, updates on disease models and interventions that modulate apoE expression and function. RESULTS: This manuscript presents highlights from the conference and provides an overview of opportunities for further research in the field. DISCUSSION: Understanding apoE's multifaceted roles in AD pathogenesis will help develop targeted interventions for AD and advance the field of AD precision medicine. HIGHLIGHTS: APOE is a central player in the pathogenesis of Alzheimer's disease. APOE exerts a numerous effects throughout the brain on amyloid-beta, tau, and other pathways. The AAIC Advancements: APOE conference encouraged discussions and collaborations on understanding the role of APOE.

Bioinformatics pipeline to guide post-GWAS studies in Alzheimer's: A new catalogue of disease candidate short structural variants.

BACKGROUND: Short structural variants (SSVs), including insertions/deletions (indels), are common in the human genome and impact disease risk. The role of SSVs in late-onset Alzheimer's disease (LOAD) has been understudied. In this study, we developed a bioinformatics pipeline of SSVs within LOAD-genome-wide association study (GWAS) regions to prioritize regulatory SSVs based on the strength of their predicted effect on transcription factor (TF) binding sites. METHODS: The pipeline utilized publicly available functional genomics data sources including candidate cis-regulatory elements (cCREs) from ENCODE and single-nucleus (sn)RNA-seq data from LOAD patient samples. RESULTS: We catalogued 1581 SSVs in candidate cCREs in LOAD GWAS regions that disrupted 737 TF sites. That included SSVs that disrupted the binding of RUNX3, SPI1, and SMAD3, within the APOE-TOMM40, SPI1, and MS4A6A LOAD regions. CONCLUSIONS: The pipeline developed here prioritized non-coding SSVs in cCREs and characterized their putative effects on TF binding. The approach integrates multiomics datasets for validation experiments using disease models.

The mechanistic role of alpha-synuclein in the nucleus: impaired nuclear function caused by familial Parkinson's disease SNCA mutations.

Alpha-synuclein SNCA has been implicated in the etiology of Parkinson's disease (PD); however, the normal function of alpha-synuclein protein and the pathway that mediates its pathogenic effect is yet to be discovered. We investigated the mechanistic role of SNCA in the nucleus utilizing isogenic human-induced pluripotent stem cells-derived neurons from PD patients with autosomal dominant mutations, A53T and SNCA-triplication, and their corresponding corrected lines by genome- and epigenome-editing. Comparisons of shape and integrity of the nuclear envelope and its resistance to stresses found that both mutations result in similar nuclear envelope perturbations that were reversed in the isogenic mutation-corrected cells. Further mechanistic studies showed that SNCA mutation has adverse effects on the nucleus by trapping Ras-related nuclear protein (RAN) and preventing it from transporting key nuclear proteins such as, DNMT3A, for maintaining normal nuclear function. For the first time, we proposed that α-syn interacts with RAN and normally functions in the nucleocytoplasmic transport while exerts its pathogenic effect by sequestering RAN. We suggest that defects in the nucleocytoplasmic transport components may be a general pathomechanistic driver of neurodegenerative diseases.

Pleiotropic Meta-Analysis of Cognition, Education, and Schizophrenia Differentiates Roles of Early Neurodevelopmental and Adult Synaptic Pathways.

Susceptibility to schizophrenia is inversely correlated with general cognitive ability at both the phenotypic and the genetic level. Paradoxically, a modest but consistent positive genetic correlation has been reported between schizophrenia and educational attainment, despite the strong positive genetic correlation between cognitive ability and educational attainment. Here we leverage published genome-wide association studies (GWASs) in cognitive ability, education, and schizophrenia to parse biological mechanisms underlying these results. Association analysis based on subsets (ASSET), a pleiotropic meta-analytic technique, allowed jointly associated loci to be identified and characterized. Specifically, we identified subsets of variants associated in the expected ("concordant") direction across all three phenotypes (i.e., greater risk for schizophrenia, lower cognitive ability, and lower educational attainment); these were contrasted with variants that demonstrated the counterintuitive ("discordant") relationship between education and schizophrenia (i.e., greater risk for schizophrenia and higher educational attainment). ASSET analysis revealed 235 independent loci associated with cognitive ability, education, and/or schizophrenia at p < 5 × 10-8. Pleiotropic analysis successfully identified more than 100 loci that were not significant in the input GWASs. Many of these have been validated by larger, more recent single-phenotype GWASs. Leveraging the joint genetic correlations of cognitive ability, education, and schizophrenia, we were able to dissociate two distinct biological mechanisms-early neurodevelopmental pathways that characterize concordant allelic variation and adulthood synaptic pruning pathways-that were linked to the paradoxical positive genetic association between education and schizophrenia. Furthermore, genetic correlation analyses revealed that these mechanisms contribute not only to the etiopathogenesis of schizophrenia but also to the broader biological dimensions implicated in both general health outcomes and psychiatric illness.

The Path to Progress Preclinical Studies of Age-Related Neurodegenerative Diseases: A Perspective on Rodent and hiPSC-Derived Models.

Alzheimer's disease (AD) and Parkinson's disease (PD) are the two most prevalent age-related neurodegenerative diseases, and currently no effective clinical treatments exist for either, despite decades of clinical trials. The failure to translate preclinical findings into effective treatments is indicative of a problem in the current evaluation pipeline for potential therapeutics. At present, there are no useful animal models for AD and PD research that reflect the entire biology of the diseases, specifically, the more common non-Mendelian forms. Whereas the field continues to seek suitable rodent models for investigating potential therapeutics for these diseases, rodent models have still been used primarily for preclinical studies. Here, we advocate for a paradigm shift toward the application of human-induced pluripotent stem cell (hiPSC)-derived systems for PD and AD modeling and the development of improved human-based models in a dish for drug discovery and preclinical assessment of therapeutic targets.

Multiplication of the SNCA locus exacerbates neuronal nuclear aging.

Human-induced Pluripotent Stem Cell (hiPSC)-derived models have advanced the study of neurodegenerative diseases, including Parkinson's disease (PD). While age is the strongest risk factor for these disorders, hiPSC-derived models represent rejuvenated neurons. We developed hiPSC-derived Aged dopaminergic and cholinergic neurons to model PD and related synucleinopathies. Our new method induces aging through a `semi-natural' process, by passaging multiple times at the Neural Precursor Cell stage, prior to final differentiation. Characterization of isogenic hiPSC-derived neurons using heterochromatin and nuclear envelope markers, as well as DNA damage and global DNA methylation, validated our age-inducing method. Next, we compared neurons derived from a patient with SNCA-triplication (SNCA-Tri) and a Control. The SNCA-Tri neurons displayed exacerbated nuclear aging, showing advanced aging signatures already at the Juvenile stage. Noteworthy, the Aged SNCA-Tri neurons showed more α-synuclein aggregates per cell versus the Juvenile. We suggest a link between the effects of aging and SNCA overexpression on neuronal nuclear architecture.

APOE: The New Frontier in the Development of a Therapeutic Target towards Precision Medicine in Late-Onset Alzheimer's.

Alzheimer's disease (AD) has a critical unmet medical need. The consensus around the amyloid cascade hypothesis has been guiding pre-clinical and clinical research to focus mainly on targeting beta-amyloid for treating AD. Nevertheless, the vast majority of the clinical trials have repeatedly failed, prompting the urgent need to refocus on other targets and shifting the paradigm of AD drug development towards precision medicine. One such emerging target is apolipoprotein E (APOE), identified nearly 30 years ago as one of the strongest and most reproduceable genetic risk factor for late-onset Alzheimer's disease (LOAD). An exploration of APOE as a new therapeutic culprit has produced some very encouraging results, proving that the protein holds promise in the context of LOAD therapies. Here, we review the strategies to target APOE based on state-of-the-art technologies such as antisense oligonucleotides, monoclonal antibodies, and gene/base editing. We discuss the potential of these initiatives in advancing the development of novel precision medicine therapies to LOAD.

Shared genetic etiology underlying late-onset Alzheimer's disease and posttraumatic stress syndrome.

INTRODUCTION: Late-onset Alzheimer's disease (LOAD) manifests comorbid neuropsychiatric symptoms and posttraumatic stress disorder (PTSD) is associated with an increased risk for dementia in late life, suggesting the two disorders may share genetic etiologies. METHODS: We performed genetic pleiotropy analysis using LOAD and PTSD genome-wide association study (GWAS) datasets from white and African-American populations, followed by functional-genomic analyses. RESULTS: We found an enrichment for LOAD across increasingly stringent levels of significance with the PTSD GWAS association (LOAD|PTSD) in the discovery and replication cohorts and a modest enrichment for the reverse conditional association (PTSD|LOAD). LOAD|PTSD association analysis identified and replicated the MS4A genes region. These genes showed similar expression pattern in brain regions affected in LOAD, and across-brain-tissue analysis identified a significant association for MS4A6A. The African-American samples showed moderate enrichment; however, no false discovery rate-significant associations. DISCUSSION: We demonstrated common genetic signatures for LOAD and PTSD and suggested immune response as a common pathway for these diseases.

Downregulation of SNCA Expression by Targeted Editing of DNA Methylation: A Potential Strategy for Precision Therapy in PD.

Elevated levels of SNCA have been implicated in the pathogenesis of Parkinson's disease (PD), while normal physiological levels of SNCA are needed to maintain neuronal function. We ought to develop new therapeutic strategies targeting the regulation of SNCA expression. DNA methylation at SNCA intron 1 regulates SNCA transcription, and PD brains showed differential methylation levels compared to controls. Thus, DNA methylation at SNCA intron 1 is an attractive target for fine-tuned downregulation of SNCA levels. Here we developed a system, comprising an all-in-one lentiviral vector, for targeted DNA methylation editing within intron 1. The system is based on CRISPR-deactivated Cas9 (dCas9) fused with the catalytic domain of DNA-methyltransferase 3A (DNMT3A). Applying the system to human induced pluripotent stem cell (hiPSC)-derived dopaminergic neurons from a PD patient with the SNCA triplication resulted in fine downregulation of SNCA mRNA and protein mediated by targeted DNA methylation at intron 1. Furthermore, the reduction in SNCA levels by the guide RNA (gRNA)-dCas9-DMNT3A system rescued disease-related cellular phenotype characteristics of the SNCA triplication hiPSC-derived dopaminergic neurons, e.g., mitochondrial ROS production and cellular viability. We established that DNA hypermethylation at SNCA intron 1 allows an effective and sufficient tight downregulation of SNCA expression levels, suggesting the potential of this target sequence combined with the CRISPR-dCas9 technology as a novel epigenetic-based therapeutic approach for PD.

Bioinformatics strategy to advance the interpretation of Alzheimer's disease GWAS discoveries: The roads from association to causation.

INTRODUCTION: Genome-wide association studies (GWAS) discovered multiple late-onset Alzheimer's disease (LOAD)-associated SNPs and inferred the genes based on proximity; however, the actual causal genes are yet to be identified. METHODS: We defined LOAD-GWAS regions by the most significantly associated SNP ±0.5 Mb and developed a bioinformatics pipeline that uses and integrates chromatin state segmentation track to map active enhancers and virtual 4C software to visualize interactions between active enhancers and gene promoters. We augmented our pipeline with biomedical and functional information. RESULTS: We applied the bioinformatics pipeline using three ∼1 Mb LOAD-GWAS loci: BIN1, PICALM, CELF1. These loci contain 10-24 genes, an average of 106 active enhancers and 80 CTCF sites. Our strategy identified all genes corresponding to the promoters that interact with the active enhancer that is closest to the LOAD-GWAS-SNP and generated a shorter list of prioritized candidate LOAD genes (5-14/loci), feasible for post-GWAS investigations of causality. DISCUSSION: Interpretation of LOAD-GWAS discoveries requires the integration of brain-specific functional genomic data sets and information related to regulatory activity.

Multi-Trait Analysis of GWAS and Biological Insights Into Cognition: A Response to Hill (2018).

Hill (Twin Research and Human Genetics, Vol. 21, 2018, 84-88) presented a critique of our recently published paper in Cell Reports entitled 'Large-Scale Cognitive GWAS Meta-Analysis Reveals Tissue-Specific Neural Expression and Potential Nootropic Drug Targets' (Lam et al., Cell Reports, Vol. 21, 2017, 2597-2613). Specifically, Hill offered several interrelated comments suggesting potential problems with our use of a new analytic method called Multi-Trait Analysis of GWAS (MTAG) (Turley et al., Nature Genetics, Vol. 50, 2018, 229-237). In this brief article, we respond to each of these concerns. Using empirical data, we conclude that our MTAG results do not suffer from 'inflation in the FDR [false discovery rate]', as suggested by Hill (Twin Research and Human Genetics, Vol. 21, 2018, 84-88), and are not 'more relevant to the genetic contributions to education than they are to the genetic contributions to intelligence'.

The effects of the TOMM40 poly-T alleles on Alzheimer's disease phenotypes.

The TOMM40 poly-T is a polymorphism in intron 6 of the TOMM40 gene, which is adjacent to and in linkage disequilibrium with APOE. Roses et al. identified the association between the length of TOMM40 poly-T with the risk and age of onset of late-onset Alzheimer's disease (LOAD). Following the original discovery, additional studies found associations between the TOMM40 poly-T and LOAD-related phenotypes independent of APOE genotypes, while others did not replicate these associations. Furthermore, the identity of the TOMM40 poly-T risk allele has been controversial between different LOAD-related phenotypes. Here, we propose a framework to address the conflicting findings with respect to the TOMM40 poly-T allele associations with LOAD phenotypes and their functional effects. The framework is used to interpret previous studies as means to gain insights regarding the nature of the risk allele, very long versus short. We suggest that the identity of the TOMM40 poly-T risk allele depends on the phenotype being evaluated, the ages of the study subjects at the time of assessment, and the context of the APOE genotypes. In concluding remarks, we outline future studies that will inform the mechanistic interpretation of the genetic data.

The effects of PPARγ on the regulation of the TOMM40-APOE-C1 genes cluster.

Chromosome 19q13.32 is a gene rich region, and has been implicated in multiple human phenotypes in adulthood including lipids traits, Alzheimer's disease, and longevity. Peroxisome Proliferator Activated Receptor Gamma (PPARγ) is a ligand-activated nuclear transcription factor that plays a role in human complex traits that are also genetically associated with the chromosome 19q13.32 region. Here, we study the effects of PPARγ on the regional expression regulation of the genes clustered within chromosome 19q13.32, specifically TOMM40, APOE, and APOC1, applying two complementary approaches. Using the short hairpin RNA (shRNA) method in the HepG2 cell-line we knocked down PPARγ expression and measured the effect on mRNA expression. We discovered PPARγ knock down increased the levels of TOMM40-, APOE-, and APOC1-mRNAs, with the highest increase in expression observed for APOE-mRNA. To complement the PPARγ knockdown findings we also examined the effects of low doses of PPARγ agonists (nM range) on mRNA expression of these genes. Low (nM) concentrations of pioglitazone (Pio) decreased transcription of TOMM40, APOE, and APOC1 genes, with the lowest mRNA levels for each gene observed at 1.5nM. Similar to the effect of PPARγ knockdown, the strongest response to pioglitazone was also observed for APOE-mRNA, and rosiglitazone (Rosi), another PPARγ agonist, produced results that were consistent with these. In conclusion, our results further established a role for PPARγ in regional transcriptional regulation of chr19q13.32, underpinning the association between PPARγ, the chr19q13.32 genes cluster, and human complex traits and disease.

Genetic analysis of α-synuclein 3' untranslated region and its corresponding microRNAs in relation to Parkinson's disease compared to dementia with Lewy bodies.

INTRODUCTION: The α-synuclein (SNCA) gene has been implicated in the etiology of Parkinson's disease (PD) and dementia with Lewy bodies (DLB). METHODS: A computational analysis of SNCA 3' untranslated region to identify potential microRNA (miRNA) binding sites and quantitative real-time polymerase chain reaction (PCR) to determine their expression in isogenic induced pluripotent stem cell-derived dopaminergic and cholinergic neurons as a model of PD and DLB, respectively, were performed. In addition, we performed a deep sequencing analysis of the SNCA 3' untranslated region of autopsy-confirmed cases of PD, DLB, and normal controls, followed by genetic association analysis of the identified variants. RESULTS: We identified four miRNA binding sites and observed a neuronal-type-specific expression profile for each miRNA in the different isogenic induced pluripotent stem cell-derived dopaminergic and cholinergic neurons. Furthermore, we found that the short structural variant rs777296100-polyT was moderately associated with DLB but not with PD. DISCUSSION: We suggest that the regulation of SNCA expression through miRNAs is neuronal-type-specific and possibly plays a part in the phenotypic heterogeneity of synucleinopathies. Furthermore, genetic variability in the SNCA gene may contribute to synucleinopathies in a pathology-specific manner.

The SSV Evaluation System: A Tool to Prioritize Short Structural Variants for Studies of Possible Regulatory and Causal Variants.

Short structural variants (SSVs) are short genomic variants (<50 bp) other than SNPs. It has been suggested that SSVs contribute to many human complex traits. However, high-throughput analysis of SSVs presents numerous technical challenges. In order to facilitate the discovery and assessment of SSVs, we have developed a prototype bioinformatics tool, "SSV evaluation system," which is a searchable, annotated database of SSVs in the human genome, with associated customizable scoring software that is used to evaluate and prioritize SSVs that are most likely to have significant biological effects and impact on disease risk. This new bioinformatics tool is a component in a larger strategy that we have been using to discover potentially important SSVs within candidate genomic regions that have been identified in genome-wide association studies, with the goal to prioritize potential functional/causal SSVs and focus the follow-up experiments on a relatively small list of strong candidate SSVs. We describe our strategy and discuss how we have used the SSV evaluation system to discover candidate causal variants related to complex neurodegenerative diseases. We present the SSV evaluation system as a powerful tool to guide genetic investigations aiming to uncover SSVs that underlie human complex diseases including neurodegenerative diseases in aging.

The genetic contributions of SNCA and LRRK2 genes to Lewy Body pathology in Alzheimer's disease.

The molecular genetic basis that leads to Lewy Body (LB) pathology in 15-20% of Alzheimer disease cases (LBV/AD) was largely unknown. Alpha-synuclein (SNCA) and Leucine-rich repeat kinase2 (LRRK2) have been implicated in the pathogenesis of Parkinson's disease (PD), the prototype of LB spectrum disorders. We tested the association of SNCA variants with LB pathology in AD. We then stratified the SNCA association analyses by LRRK2 genotype. We also investigated the expression regulation of SNCA and LRRK2 in relation to LB pathology. We evaluated the differences in SNCA-mRNA and LRRK2-mRNA levels as a function of LB pathology in the temporal cortex (TC) from autopsy-confirmed LBV/AD cases and AD controls. We further investigated the cis-effect of the LB pathology-associated genetic variants within the SNCA and LRRK2 loci on the mRNA expression of these genes. SNCA SNPs rs3857059 and rs2583988 showed significant associations with increased risk for LB pathology. When the analyses were stratified by LRRK2-rs1491923 genotype, the associations became stronger for both SNPs and an association was also observed with rs2619363. Expression analysis demonstrated that SNCA- and LRRK2-mRNA levels were significantly higher in TC from LBV/AD brains compared with AD controls. Furthermore, SNCA-mRNA expression level in the TC was associated with rs3857059; homozygotes for the minor allele showed significant higher expression. LRRK2-transcript levels were increased in carriers of rs1491923 minor allele. Our findings demonstrated that SNCA contributes to LB pathology in AD patients, possibly via interaction with LRRK2, and suggested that expression regulation of these genes may be the molecular basis underlying the observed LB associations.

A cytosine-thymine (CT)-rich haplotype in intron 4 of SNCA confers risk for Lewy body pathology in Alzheimer's disease and affects SNCA expression.

INTRODUCTION: We recently showed that tagging single-nucleotide polymorphisms across the SNCA locus were significantly associated with increased risk for Lewy body (LB) pathology in Alzheimer's disease (AD) cases. However, the actual genetic variant(s) that underlie the observed associations remain elusive. METHODS: We used a bioinformatics algorithm to catalog structural variants in a region of SNCA intron 4, followed by phased sequencing. We performed a genetic association analysis in autopsy series of LB variant of Alzheimer's disease (LBV/AD) cases compared with AD-only controls. We investigated the biological functions by expression analysis using temporal-cortex samples. RESULTS: We identified four distinct haplotypes within a highly polymorphic low-complexity cytosine-thymine (CT)-rich region. We showed that a specific haplotype conferred risk to develop LBV/AD. We demonstrated that the CT-rich site acts as an enhancer element, where the risk haplotype was significantly associated with elevated levels of SNCA messenger RNA. DISCUSSION: We have discovered a novel haplotype in a CT-rich region in SNCA that contributes to LB pathology in AD patients, possibly via cis-regulation of the gene expression.

The cis-regulatory effect of an Alzheimer's disease-associated poly-T locus on expression of TOMM40 and apolipoprotein E genes.

BACKGROUND: We investigated the genomic region spanning the Translocase of the Outer Mitochondrial Membrane 40-kD (TOMM40) and Apolipoprotein E (APOE) genes, that has been associated with the risk and age of onset of late-onset Alzheimer's disease (LOAD) to determine whether a highly polymorphic, intronic poly-T within this region (rs10524523; hereafter, 523) affects expression of the APOE and TOMM40 genes. Alleles of this locus are classified as S, short; L, long; and VL, very long based on the number of T residues. METHODS: We evaluated differences in APOE messenger RNA (mRNA) and TOMM40 mRNA levels as a function of the 523 genotype in two brain regions from APOE ε3/ε3 white autopsy-confirmed LOAD cases and normal controls. We further investigated the effect of the 523 locus in its native genomic context using a luciferase expression system. RESULTS: The expression of both genes was significantly increased with disease. Mean expression of APOE and TOMM40 mRNA levels were higher in VL homozygotes compared with S homozygotes in the temporal and occipital cortexes from normal and LOAD cases. Results of a luciferase reporter system were consistent with the human brain mRNA analysis; the 523 VL poly-T resulted in significantly higher expression than the S poly-T. Although the effect of poly-T length on reporter expression was the same in HepG2 hepatoma and SH-SY5Y neuroblastoma cells, the magnitude of the effect was greater in the neuroblastoma than in the hepatoma cells, which implies tissue-specific modulation of the 523 poly-T. CONCLUSIONS: These results suggest that the 523 locus may contribute to LOAD susceptibility by modulating the expression of TOMM40 and/or APOE transcription.

African-American TOMM40'523-APOE haplotypes are admixture of West African and Caucasian alleles.

BACKGROUND: Several studies have demonstrated a lower apolipoprotein E4 (APOE ε4) allele frequency in African-Americans, but yet an increased age-related prevalence of AD. An algorithm for prevention clinical trials incorporating TOMM40'523 (Translocase of Outer Mitochondria Membrane) and APOE depends on accurate TOMM40'523-APOE haplotypes. METHODS: We have compared the APOE and TOMM40'523 phased haplotype frequencies of a 9.5 kb TOMM40/APOE genomic region in West African, Caucasian, and African-American cohorts. RESULTS: African-American haplotype frequency scans of poly-T lengths connected in phase with either APOE ε4 or APOE ε3 differ from both West Africans and Caucasians and represent admixture of several distinct West African and Caucasian haplotypes. A new West African TOMM40'523 haplotype, with APOE ε4 connected to a short TOMM40'523 allele, is observed in African-Americans but not Caucasians. CONCLUSION: These data have therapeutic implications for the age of onset risk algorithm estimates and the design of a prevention trial for African-Americans or other mixed ethnic populations.

Neuronal-type-specific epigenome editing to decrease SNCA expression: Implications for precision medicine in synucleinopathies.

Overexpression of SNCA has been implicated in the pathogenesis of synucleinopathies, particularly Parkinson's disease (PD) and dementia with Lewy bodies (DLB). While PD and DLB share some clinical and pathological similarities, each disease presents distinct characteristics, including the primary affected brain region and neuronal type. We aimed to develop neuronal-type-specific SNCA-targeted epigenome therapies for synucleinopathies. The system is based on an all-in-one lentiviral vector comprised of CRISPR-dSaCas9 and guide RNA (gRNA) targeted at SNCA intron 1 fused with a synthetic repressor molecule of Krüppel-associated box (KRAB)/ methyl CpG binding protein 2 (MeCp2) transcription repression domain (TRD). To achieve neuronal-type specificity for dopaminergic and cholinergic neurons, the system was driven by tyrosine hydroxylase (TH) and choline acetyltransferase (ChAT) promoters, respectively. Delivering the system into human induced pluripotent stem cell (hiPSC)-derived dopaminergic and cholinergic neurons from a patient with the SNCA triplication resulted in efficient and neuronal-type-specific downregulation of SNCA-mRNA and protein. Furthermore, the reduction in SNCA levels by the gRNA-dSaCas9-repressor system rescued disease-related cellular phenotypes including Ser129-phophorylated α-synuclein, neuronal viability, and mitochondrial dysfunction. We established a novel neuronal-type-specific SNCA-targeted epigenome therapy and provided in vitro proof of concept using human-based disease models. Our results support the therapeutic potential of our system for PD and DLB and provide the foundation for further preclinical studies in animal models toward investigational new drug (IND) enablement and clinical trials.