iPSC-based Drug Screening for PARK9, a Familial Parkinson’s Disease with Impaired Autophagy

Abstract

Parkinson's disease (PD) is a neurodegenerative disorder with the degeneration of midbrain dopaminergic neurons. However, few disease-modifying drugs for patients with PD have been identified. Familial Parkinson's diseases are numbered in the order in which the causative genes are identified, and one of them, PARK9, is caused by autophagy dysfunction and has mutation in the ATP13A2 gene which encodes a lysosomal type 5 P-type ATPase. To elucidate the pathogenesis and to screen effective drugs for PARK9, we generated iPSCs from the T cells from a PARK9 patient. Three independent clones were used for the analysis after quality verification. First, we confirmed reduced autophagy function in dopaminergic neurons derived from PARK9 iPSCs by examining the accumulation of LC3B, an autophagosome marker, due to impaired lysosomal functions. Then, we successfully quantified LC3B accumulation in the cell bodies of PARK9 dopaminergic neurons using IN Cell Analyzer 2200. Using these methods, we screened a compound library containing 320 chemical compounds using reduced LC3B accumulation in PARK9 dopaminergic neurons as an indicator. The first screening narrowed the list of candidate compounds to 70, followed by the second screening to exclude the compounds that reduce LC3B production. This two-step and 96 well-based high-throughput screening resulted in the identification of five novel disease modifying drug candidates that ameliorate impaired lysosomal dysfunction in PARK9-dopaminergic neurons.