Concise synthesis and biological activity evaluation of novel pyrazinyl–aryl urea derivatives against several cancer cell lines, which can especially induce T24 apoptotic and necroptotic cell death

Abstract

Based on the structural modification of regorafenib, 28 pyrazinyl–aryl urea derivatives were synthesized and their in vitro antiproliferative activities were evaluated. Six compounds (5-16, 5-17, 5-18, 5-19, 5-22, and 5-23) exhibited favorable inhibitory activity against the human bladder cancer T24 cell line, and 5-23 demonstrated the strongest inhibitory activity (IC₅₀ = 4.58 ± 0.24 μM) with high selectivity. Compound 5-23 induced apoptosis in the low concentration range (≤7.5 μM) combined with shorter incubation time (≤10 h) via the activation of caspases, while high concentrations and prolonged incubation times led to necroptotic cell death by activating the RIPK1/RIPK3/MLKL signaling pathway. Induced apoptosis and necroptosis were closely associated with intracellular reactive oxygen species generation and decreased mitochondrial membrane potential. Compared with regorafenib, 5-23 displayed improved pharmacokinetic profiles in an in vivo rat model. Molecular docking and structure–activity relationship analyses were in agreement with the biological data. Compound 5-23 may be a potent anti-bladder cancer agent and this small molecule can be considered as a promising structure for further optimization.