Discovery of indolinone-based covalent ULK1 inhibitors that suppressed autophagy and induced apoptosis against colorectal carcinoma.

Unc-51 Like Autophagy Activating Kinase 1 (ULK1) is an essential regulator in the initiation of autophagy and represents a novel therapeutic target for colorectal cancer (CRC) treatment. Herein, we developed a series of novel ULK1 covalent inhibitors through structure-based and medicinal chemistry optimizations. Notably, compound 12i was synthesized and validated to form covalent bonds with ULK1, exhibiting pronounced selectivity and potent inhibitory activity (ULK1 IC50 = 4.7 μM). Moreover, 12i demonstrated anti-proliferative effects in CRC cell lines. In vitro experiments showed that 12i effectively inhibited CRC cell growth, induced cell cycle arrest at the S or G2/M phase, and triggered apoptosis by blocking ULK1-mediated autophagy. Additionally, in vivo results showed that 12i also exhibited good tumor inhibitory activity in CRC murine models, with an inhibition rate of 70.4 % at 40 mg/kg. In vivo pharmacokinetic studies revealed moderate systemic absorption (Tmax = 2 h) with oral bioavailability of 39.7 % in rats. In summary, our findings highlight compound 12i as a promising lead candidate for further development of selective ULK1 covalent inhibitors, offering a potential new strategy for CRC therapy.