Liver cancer is the second cause of death from cancer worldwide, without effective treatment. Traditional chemotherapy for liver cancer has big side effects for patients, whereas targeted drugs, such as sorafenib, commonly have drug resistance. Oroxylin A (OA) is the main bioactive flavonoids of Scutellariae radix, which has strong anti-hepatoma effect but low toxicity to normal tissue. To date, no differentiation-inducing agents have been reported to exert a curative effect on solid tumors. Here our results demonstrated that OA restrained the proliferation and induced differentiation of hepatoma both in vitro and in vivo, via inducing a high PKM1 (pyruvate kinase M1)/PKM2 (pyruvate kinase M2) ratio. In addition, inhibited expression of polypyrimidine tract-binding protein by OA was in charge of the decrease of PKM2 and increase of PKM1. Further studies demonstrated that increased PKM1 translocated into the nucleus and bound with HNF-4α (hepatocyte nuclear factor 4 alpha) directly, promoting the transcription of HNF-4α-targeted genes. This work suggested that OA increased PKM1/PKM2 ratio, resulting in HNF-4α activation and hepatoma differentiation. Especially, OA showed reliable anticancer effect on both human primary hepatocellular carcinoma cells and patient-derived tumor xenograft model for hepatoma, and slowed down the development of primary hepatoma, suggesting that OA could be developed into a novel differentiation inducer agent for hepatoma.