Restoring Parkin Function: An AAV Gene Therapy Approach for Early-Onset Parkinson's Disease.

BACKGROUND

Biallelic loss-of-function mutations in PRKN gene (encoding Parkin protein) cause early-onset Parkinson's disease (EOPD). Parkin is a crucial component of PINK1-Parkin pathway, which marks damaged mitochondria for degradation via mitophagy. Without functional Parkin, damaged mitochondria accumulate, causing oxidative stress and neurodegeneration.

OBJECTIVE

Investigate Parkin gene replacement via AAV gene therapy as a potential treatment for Parkin-dependent EOPD.

METHODS

We initially validated phosphorylated ubiquitin Ser65 (pUb Ser65 ) as an indicator of Parkin-mediated mitophagy initiation. We evaluated AAV-mediated PRKN replacement (hereafter, AAV-Parkin) in a Parkin knockout neuroblastoma cell line (SH-SY5Y cells) and feasibility of delivery in mouse and rat models.

RESULTS

Our research showed pUb Ser65 signal was reduced in Parkin-KO SH-SY5Y cells when compared to wild-type cells after mitochondrial stress, indicating deficiency in initiation of mitophagy. AAV-mediated human PRKN gene replacement successfully restored these pUb Ser65 levels in knockout cells. We saw restoration in patient-derived fibroblasts following AAV-Parkin overexpression. We developed a translatable gene therapy approach using rodents. We demonstrated the feasibility of delivering AAV-Parkin directly into the substantia nigra (SN) of wild-type rats. Using an AAV1 capsid with Ef1a promoter, we achieved dose-dependent Parkin expression and identified a well-tolerated dose. We also evaluated multiple promoters in a proprietary Spark100 capsid, finding Ef1a and Synapsin1 (Syn1) were most effective for transducing dopaminergic neurons in the SN of mice without causing adverse effects. These findings established a well-tolerated vector dose and an optimal capsid-promoter combination.

CONCLUSIONS

Our results support the potential of AAV-Parkin gene therapy as a disease-modifying approach for Parkin-deficient EOPD.