Off-the-shelf Bioreactor Produced, iPSC-derived Neural Microtissues Containing Dopaminergic Neurons Innervate the Striatum and Normalize Behavior in a Parkinson Rat Model
Nicolas Prudon1, Lucía Cordero-Espinoza1, Chloé Morel1, Marilyn Lepleux1, Basile Gurchenkov1, Myriam Abarkan1, Loanne Milvoy1, Pauline Morand1, Fabien Moncaubeig1, Hélène Wurtz1, Léa Poinçot1, Maëlle Demarco1, Agathe Jonckeau1, Justine Plétenka1, Elisa Luquet1, Michael Lanero Fidalgo1, Guillaume Dabee1, Thibault Dufourd1, Jens Schroeder1, Kevin Alessandri1, Maxime Feyeux1, Erwan Bezard2, Emilie Faggiani1
1Treefrog Therapeutics, 2Institut des Maladies Neurodégénératives (CNRS UMR 5293), Université de Bordeaux
Objective:
To demonstrate the efficacy of a ready-to-graft 3D neural microtissue product – manufactured at large scale - as a therapeutically viable option to treat Parkinson’s disease.
Background:
A breadth of preclinical studies (Doi et al, 2020; Piao et al, 2021; Hiller et al, 2022, Kirkeby et al, 2023) is now backing the rationale of pluripotent-stem cell (PSC)-derived cell replacement therapies to restore motor function in Parkinsonian patients. The target for replacement is the major dysfunctional cell population in the disease: ventral mesencephalic A9 dopaminergic neurons, which are particularly vulnerable to the in vitro manipulations required for intracerebral administration (Marchionini et al., 2003). Optimizing for survival and functionality post-transplantation is thus a significant pharmaceutical hurdle for manufacturing and delivering dopaminergic neuron-containing cell therapies. We report here a change in graft format, i.e. moving away from cell suspensions to 3D neural microtissues that are resistant to the stress of transplantation.
Design/Methods:
We utilized TreeFrog Therapeutics’ proprietary high-throughput cell-encapsulation technology (C-stem™) and standard bioreactors to provide a biomimetic 3D culture environment that enables the growth factor-directed differentiation of iPSCs into neural microtissues fit-for-cryopreservation. The product was characterized using orthogonal methods including flow cytometry, immunofluorescence labelling, RTqPCR and bulkRNAseq. Controlled doses of microtissues were administered into the striatum of 6-OHDA lesioned nude rats for functional assessment.
Results:
We demonstrate a scalable process to generate off-the-shelf cryopreserved iPSC-derived 3D neural microtissues containing a mixture of ventral mesencephalic dopaminergic neurons and dopaminergic progenitors. Upon administration, the neural microtissues innervate the lesioned striatum of hemiparkinsonian rodents with TH+ dopaminergic projections and lead to motor recovery by 16 weeks (MFD and high dose) and 20 weeks (low dose) respectively.
Conclusions:
These data establish proof-of-concept efficacy of the dopaminergic neuron-containing neural microtissue product and support the intention to pursue preclinical studies to assess its safety and efficacy as a cell therapy for Parkinson's disease.