Search

Human pluripotent stem cells show considerable promise for applications in regenerative medicine, including the development of cell replacement paradigms for the treatment of Parkinson's disease. Protocols have been developed to generate authentic midbrain dopamine (mDA) neurons capable of reversing dopamine-related deficits in animal models of Parkinson's disease. However, the generation of mDA neurons at clinical scale suitable for human application remains an important challenge. Here, we present an mDA neuron derivation protocol based on a two-step WNT signaling activation strategy that improves expression of midbrain markers, such as Engrailed-1 (EN1), while minimizing expression of contaminating posterior (hindbrain) and anterior (diencephalic) lineage markers. The resulting neurons exhibit molecular, biochemical, and electrophysiological properties of mDA neurons. Cryopreserved mDA neuron precursors can be successfully transplanted into 6-hydroxydopamine (6OHDA) lesioned rats to induce recovery of amphetamine-induced rotation behavior. The protocol presented here is the basis for clinical-grade mDA neuron production and preclinical safety and efficacy studies., Competing Interests: Declaration of Interests L.S. is a scientific co-founder and consultant, and V.T. is a founding investigator and consultant, of Bluerock Therapeutics, Inc. M.T. and S.I. are employed by BlueRock Therapeutics, Inc. L.S., S.K., S.I., and M.T. are inventors of a patent WO2016196661A1 filed by Memorial Sloan Kettering Cancer Center on the methods described in this study. All other authors declare no competing interests., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Parkinson's disease is characterized by the loss of dopaminergic neurons in the substantia nigra leading to disabling deficits. Dopamine neuron grafts may provide a significant therapeutic advance over current therapies. We have generated midbrain dopamine neurons from human embryonic stem cells and manufactured large-scale cryopreserved dopamine progenitors for clinical use. After optimizing cell survival and phenotypes in short-term studies, the cell product, MSK-DA01, was subjected to an extensive set of biodistribution, toxicity, and tumorigenicity assessments in mice under GLP conditions. A large-scale efficacy study was also performed in rats with the same lot of cells intended for potential human use and demonstrated survival of the grafted cells and behavioral amelioration in 6-hydroxydopamine lesioned rats. There were no adverse effects attributable to the grafted cells, no obvious distribution outside the brain, and no cell overgrowth or tumor formation, thus paving the way for a future clinical trial., Competing Interests: Declaration of Interests V.T. and L.S. are scientific co-founders and consultants of BlueRock Therapeutics. M.T., S.I., and S.M. are currently employed at BlueRock Therapeutics Inc. Other authors declare no competing interests. MSK-DA01 is the subject of a patent, US2018/0094242 A1, filed by the Memorial Sloan Kettering Cancer Center., (Copyright © 2021 Elsevier Inc. All rights reserved.)