Discovery of novel aniline-pyrimidine analogues as orally bioavailable dual VEGFR-2/tubulin inhibitors with potent antitumor and anti-angiogenesis efficacy.

Combinations of anti-angiogenesis agents with microtubule-targeting drugs have shown promise in clinical cancer therapy. Considering the advantages of dual-targeting agents over combination regimens, we designed and synthesized a series of novel small molecules capable of concurrently inhibiting VEGFR-2 and tubulin. Among these, compound 19d emerged as a highly potent candidate, exhibiting low cytotoxicity along with robust inhibitory activity against both VEGFR-2 and tubulin. It also displayed superior anti-proliferative effects across a panel of cancer cell lines, including those with drug-resistant phenotypes. Mechanistic investigations indicated that 19d exerted its antitumor effects through multifaceted pathways, including enhancing reactive oxygen species (ROS) generation, disrupting mitochondrial membrane potential, inducing apoptosis, and arresting cell cycle progression. Moreover, 19d exhibited pronounced anti-angiogenic properties, significantly impairing human umbilical vein endothelial cell (HUVEC) migration, invasion, and tube formation in vitro. Furthermore, compound 19d also demonstrated favorable metabolic stability and high oral bioavailability. Finally, in a zebrafish xenograft model, 19d potently suppressed angiogenesis, tumor growth, and metastasis. Collectively, these findings underscore the potential of 19d as a promising lead compound for the development of dual-target anti-cancer therapeutics.