Date Published: November 20, 2018
Publisher: John Wiley and Sons Inc.
Author(s): Silvia Bolognin, Marie Fossépré, Xiaobing Qing, Javier Jarazo, Janez Ščančar, Edinson Lucumi Moreno, Sarah L. Nickels, Kobi Wasner, Nassima Ouzren, Jonas Walter, Anne Grünewald, Enrico Glaab, Luis Salamanca, Ronan M. T. Fleming, Paul M. A. Antony, Jens C. Schwamborn.
Parkinson’s disease (PD)‐specific neurons, grown in standard 2D cultures, typically only display weak endophenotypes. The cultivation of PD patient‐specific neurons, derived from induced pluripotent stem cells carrying the LRRK2‐G2019S mutation, is optimized in 3D microfluidics. The automated image analysis algorithms are implemented to enable pharmacophenomics in disease‐relevant conditions. In contrast to 2D cultures, this 3D approach reveals robust endophenotypes. High‐content imaging data show decreased dopaminergic differentiation and branching complexity, altered mitochondrial morphology, and increased cell death in LRRK2‐G2019S neurons compared to isogenic lines without using stressor agents. Treatment with the LRRK2 inhibitor 2 (Inh2) rescues LRRK2‐G2019S‐dependent dopaminergic phenotypes. Strikingly, a holistic analysis of all studied features shows that the genetic background of the PD patients, and not the LRRK2‐G2019S mutation, constitutes the strongest contribution to the phenotypes. These data support the use of advanced in vitro models for future patient stratification and personalized drug development.
The identification of promising drug candidates in preclinical research, as well as personalized precision medicine, is hampered by the lack of sufficiently representative in vitro models. This is particularly true in the case of Parkinson’s disease (PD), a complex neurodegenerative disorder where the most studied cells, associated to the onset of motor dysfunctions, are the dopaminergic neurons of the substantia nigra in the midbrain.1 PD is a disorder for which animal models are not sufficiently predictive of the human response. The difficulty of capturing the multifactorial nature of the disease in conventional in vitro models and the excessive reliance on animal models partly explain the disappointingly high failure rate of new candidate molecules in clinical trials.1, 2 New technological advancements have failed to translate into successful curative pharmacological options and no definitive disease‐modifying therapy is currently available.
Recapitulating the key cellular hallmarks of LRRK2‐associated toxicity in patient‐derived cells is a prerequirement to set up in vitro assays, which can drive personalized medicine approaches. Here, we developed a platform based on hNESC‐derived neurons from PD patients carrying the LRRK2‐G2019S mutation. This platform recapitulates key features of PD, including degeneration of dopaminergic neurons and preceding mitochondrial impairments. Treatment with the LRRK2‐specific inhibitor, Inh2, rescued neurodegeneration, and neurite complexity phenotypes without fully reversing mitochondrial abnormalities. When considering all experiments at all time points, genetic background of the patients was found to be a major discriminating factor among the lines and not the LRRK2‐G2019S mutation.
Cell Lines: In this paper, the cell lines used for all the experiments, unless otherwise indicated, are described in Figure S1A (Supporting Information). Two lines were obtained from two healthy individuals and two lines from two PD patients carrying the LRRK2‐G2019S mutation. To create the isogenic lines, the mutation was either corrected or introduced. From Figures 1 to 3 and in the Supporting Information, the grouping for the lines was done according to the LRRK2 sequence (LRRK2‐WT or LRRK2‐G2019S), independently of the genetic background (healthy or PD). Only in Figure 4, the genetic background of the lines was taken into account. In Figure S3A,B (Supporting Information), the microarray analysis was performed in hNESC from six healthy individuals and six PD patients carrying LRRK2‐G2019S.
The authors declare no conflict of interest.