Ovarian cancer is associated with a high percentage of recurrence of tumors and resistance to chemotherapy. Cancer stem cells (CSCs) are responsible for cancer progression, tumor recurrence, metastasis, and chemoresistance. Thus, developing CSC-targeting therapy is an urgent need in cancer research and clinical application. In an attempt to achieve potent and selective anti-CSC agents, a series of celastrol derivatives with cinnamamide chains were synthesized and evaluated for their anti-ovarian cancer activities. Most of the compounds exhibited stronger antiproliferative activity than celastrol, and celastrol derivative 7g with a 3,4,5-trimethoxycinnamamide side chain was found to be the most potent antiproliferative agent against ovarian cancer cells with an IC<sub>50</sub> value of 0.6 μM. Additionally, compound 7g significantly inhibited the colony formation ability and reduced the number of tumor spheres. Furthermore, compound 7g decreased the percentage of CD44<sup>+</sup>, CD133<sup>+</sup> and ALDH<sup>+</sup> cells. Thus, compound 7g is a promising anti-CSC agent and could serve as a candidate for the development of new anti-ovarian cancer drugs.