A series of novel quinoline-chalcone derivatives were designed, synthesized, and evaluated for their antiproliferative activity. Among them, compound 24d exhibited the most potent activity with IC<sub>50</sub> values ranging from 0.009 to 0.016 μM in a panel of cancer cell lines. Compound 24d also displayed a good safety profile with an LD<sub>50</sub> value of 665.62 mg/kg by intravenous injection, and its hydrochloride salt 24d-HCl significantly inhibited tumor growth in H22 xenograft models without observable toxic effects, which was more potent than that of CA-4. Mechanism studies demonstrated that 24d bound to the colchicine site of tubulin, arrested the cell cycle at the G2/M phase, induced apoptosis, depolarized mitochondria, and induced reactive oxidative stress generation in K562 cells. Moreover, 24d has potent in vitro antimetastasis and in vitro and in vivo antivascular activities. Collectively, our findings suggest that 24d deserves to be further investigated as a potent and safe antitumor agent for cancer therapy.