ANALYSIS OF NEURODEVELOPMENTAL DEFECTS IN HUMAN MIDBRAIN ORGANOIDS FROM GBA-N370S PARKINSON'S DISEASE PATIENTS

With increasing prevalence, Parkinson’s disease presents a major challenge for medical research
and public health. Despite years of investigation, significant knowledge gaps exist and Parkinson’s
disease (PD) etiology remains unclear. A recent concept in the field is that neurodevelopmental
aspects might contribute to the pathogenesis of neurodegenerative diseases such as PD.
Our hypothesis is that mutations in PD-linked genes have an impact on the cells’ homeostasis at
the neural precursor state, giving rise to vulnerable dopaminergic (DA) neurons, thereby increasing
the degree of susceptibility for neurodegeneration with aging. In order to investigate this, we used
a human midbrain organoid (hMO) model generated from iPSC-derived neural precursor cells. As
part of the optimization of the model, we treated the organoids with the neurotoxin 6-OHDA to
develop a neurotoxin-induced PD model and set up a high-content imaging pipeline coupled with
machine learning classification to predict neurotoxicity. We then used these tools to derive PD
patient-derived hMOs in order to investigate our main hypothesis. First, we focused on PD patients
carrying a heterozygous mutation in the GBA gene. We developed a genome-scale metabolic
model that predicted significant differences in lipid metabolism between patients and controls.
Then, we validated the observations by performing a comprehensive lipidomics analysis confirming
a dysregulated lipidome in mutant hMOs. Moreover, GBA-PD hMOs displayed PD-relevant
phenotypes, impaired DA differentiation and an increased population of neural progenitor cells
(NPCs) in cell cycle arrest, confirming the presence of neurodevelopmental defects. To further
investigate the neurodevelopmental component of PD, we used patient-derived cell lines carrying
PINK1 mutations. PINK1-PD neural precursors presented differences in their energetic profile,
imbalanced proliferation, apoptosis, mitophagy, and an impaired differentiation efficiency to DA
neurons compared to controls. Correction of the PINK1 point mutation was able to improve the
metabolic properties and neuronal firing rates as well as rescuing the differentiation phenotype.
We performed a drug screen using repurposed drugs as well as novel compounds to evaluate their
potential to rescue the observed developmental phenotype. Treatment with 2-hydroxypropyl-β-
cyclodextrin increased the autophagy and mitophagy capacity of neurons which was accompanied
by improved dopaminergic differentiation of patient-specific neurons in midbrain organoids and
showed neuroprotective effects in an MPTP-treated mice PD model.
In conlusion, PD has a neurodevelopmental component that increases susceptibility to the
pathology. Thus, our findings suggest that the use of hMOs are suitable to reveal early PD
pathomechanisms, as well as constituting a powerful tool for advanced therapy development.