AbstractHuman urine cells (HUCs) can be reprogrammed into neural progenitor cells (NPCs) or induced pluripotent stem cells (iPSCs) with defined factors and a small molecule cocktail, but the underlying fate choice remains unresolved. Here, through sequential removal of individual compound from small molecule cocktail, we showed that A8301, a TGFβ signaling inhibitor, is sufficient to switch the cell fate from iPSCs into NPCs in OSKM-mediated HUCs reprogramming. However, TGFβ exposure at early stage inhibits HUCs reprogramming by promoting EMT. Base on these data, we developed an optimized approach for generation of NPCs or iPSCs from HUCs with significantly improved efficiency by regulating TGFβ activity at different reprogramming stages. This approach provides a simplified and improved way for HUCs reprogramming, thus would be valuable for banking human iPSCs or NPCs from people with different genetic background.