Abstract Body

Aims: Elevated SNCA levels are causative in Parkinson’s Disease (PD) pathogenesis. Patients with SNCA triplication and duplication suffer from familial early onset PD, suggesting a therapeutics window of <30%. We aim to translate mechanistic knowledge of SNCA dysregulation towards the development of epigenome therapy targeting SNCA expression.

Methods: We developed all-in-one lentiviral vector carrying the CRISPR/deactivated(d)Cas9 and selected gRNAs targeted at SNCA-intron1 fused with effector molecules specifically, the catalytic domain of DNMT3A or the transcription repressors KRAB/MeCP2.

Results: We previously transduced the gRNA-dCas9-repressor system into human iPSC-derived ‘aged’ dopaminergic neurons from a PD-patient with the SNCA triplication and showed fine-tuned downregulation of SNCA-mRNA and protein levels that led to the rescue of disease-related pathalogical phenotypes including, mitochondrial dysfunction, neuronal-cell death, DNA damage and nuclear deficits. We now pursued with in vivo validation studies. Stereotaxic injection of the gRNA/dCas9-effector system into the mouse sub-nigra lowered Snca-mRNA and protein expression levels, whereas the effect of the KRAB-MeCP2 construct was greater than that of the DNMT3A. The effect on gene expression was specific to Snca gene and no changes were observed in other brain structures including the striatum and cerebellum. Safety measures demonstrated no abnormalities in daily monitored well-being criterions and no weight loss.

Conclusions: We developed a novel technology to intervene with the transcription program of SNCA, based on precise epigenome editing, and validated effective and specific fine-tuned reduction in SNCA expression sufficient for reversing PD-associated perturbations. This study provides a proof-of-concept for advancing the development towards precision medicine for PD.