Thieno[2,3-d]pyrimidine scaffold optimization for synergistic c-Met/STAT3 blockade: Synthesis, structure-activity relationships, in vitro potency, and apoptotic pathways in colorectal cancer.

Abstract

A series of thieno[2,3-d]pyrimidine derivatives was rationally designed based on our previously reported pyrazolo[3,4-d]pyrimidine scaffold as dual c-Met/STAT3 inhibitors to block both related oncogenic signaling pathways. Among the synthesized derivatives, compounds 17d and 17e were found to be the most potent and exhibited significant anticancer activity across a wide range of cancer cell lines within the NCI-60 panel. Both derivatives demonstrated remarkable anticancer efficacy and selectivity toward HCT-116 colorectal cancer cells, with better cytotoxicity than the clinical drug doxorubicin. Remarkably, compounds 17d and 17e demonstrated potent inhibitory activity against two targets, c-Met and STAT3, with submicromolar IC50 values compared to the reference inhibitors. According to the flow cytometry data, both compounds induced G2/M cell cycle arrest. While both compounds 16d and 17d induced apoptosis in HCT-116 cells, compound 17d additionally triggered necrotic cell death. In addition, compound 17e caused a promising pro-apoptotic transcriptional by upregulation of p53, Bax, PUMA, and caspases-3/8/9 alongside Bcl-2 downregulation in HCT-116 cells. The molecular docking studies confirmed the proposed mechanism of action and proved that the designed molecules bind efficiently to the active site of both protein targets. Finally, compounds 17d and 17e were found to possess good drug-likeness, and the predicted ADMET properties were within an acceptable range. Thus, these derivatives are promising leads for further development of dual c-Met/STAT3 inhibitors for treating colorectal cancer.