Palladium-catalyzed C(sp3)-H arylation of celastrol: synthesis and pharmacological evaluation of C-21/28 arylated derivatives

Celastrol is a quinone methide triterpenoid with potent antitumor activity, yet its structural modifications are mostly limited to the C-29 carboxyl group, leaving Other sites underexplored. To expand its structural diversity, we employed the well-established palladium-catalyzed C(sp³)-H arylation strategy to prepare a series of novel arylated derivatives at the C-21/28 positions of the E-ring. Their structures were fully confirmed by 2D-NMR and HRMS. Most of these derivatives displayed potent anti-proliferative effects against Cancer cells. Among them, 28a showed the strongest activity against HCT116 cells (IC₅₀ = 0.34 μM), which was superior to that of celastrol (IC₅₀ = 0.82 μM). Notably, 28a showed greatly improved tumor selectivity, with a selectivity index 6-8-fold higher than celastrol in normal human cells L02 and LX2. Mechanistically, 28a triggered ROS accumulation, mitochondrial membrane potential depolarization, Apoptosis, and cell cycle arrest in HCT116 cells, accompanied by downregulation of Bcl-2/Bcl-xL and upregulation of γ-H2AX. In acute toxicity tests, 28a showed a more favorable safety profile than celastrol in mice. In a colorectal tumor xenograft model, 28a displayed significant antitumor efficacy (TGI = 47.9%). This work demonstrates that E-ring C(sp³)-H arylation is an effective strategy for the modification of celastrol, providing valuable lead compounds for further structural optimization and antitumor activity evaluation.