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.

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.

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.  

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.

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.