Direct conversion generates patient-specific, disease-relevant cell types, such as neurons, that are rare, limited, or difficult to isolate from common and easily accessible cells, such as skin cells. However, low rates of direct conversion and complex protocols limit scalability and, thus, the potential of cell-fate conversion for biomedical applications. Here, we optimize the conversion protocol by examining process parameters, including transcript design
delivery via adeno-associated virus (AAV), retrovirus, and lentivirus
cell seeding density
and the impact of media conditions. Thus, we report a compact, portable conversion process that boosts proliferation and increases direct conversion of mouse fibroblasts to induced motor neurons (iMNs) to achieve high conversion rates of above 1,000%, corresponding to more than ten motor neurons yielded per cell seeded, which we achieve through expansion. Our optimized, direct conversion process generates functional motor neurons at scales relevant for cell therapies (>
10