Design, Synthesis and in Vitro Tumor Cytotoxicity Evaluation of 3,5-Diamino-N-substituted Benzamide Derivatives as Novel GSK-3β Small Molecule Inhibitors

Abstract

Glycogen synthase kinase-3 (GSK-3) plays an important regulatory role in various signaling pathways; such as PI3 K/AKT, which is closely related to the occurrence and development of tumors. At present, the most reported active GSK-3 inhibitors have the same structure: lactam ring or amide structure. To find out the GSK-3β small molecule inhibitor with novel, safe, efficient and more uncomplicated synthesis method, we analyzed in-depth reported crystal-binding patterns of GSK-3β small molecule inhibitor with GSK-3β protein, and designed and synthesized 17 non-reported 3,5-diamino-N-substituted benzamide compounds. Their structures were confirmed by ¹ H-NMR, ¹³ C-NMR, and HR-MS. The preliminary screening of tumor cytotoxicity of compounds in vitro was detected by MTT, and their structure-activity relationships were illustrated. The results have shown that 3,5-diamino-N-[3-(trifluoromethyl)phenyl]benzamide (4d) exhibited significant tumor cytotoxicity against human colon cancer cells (HCT-116) with IC₅₀ of 8.3 μm and showed commendable selectivity to GSK-3β. In addition, Compound 4d induced apoptosis to some extent and possessed modest PK properties.

Keywords: 3,5-diamino-N-substituted benzamide; GSK-3β; cytotoxicity; small molecule inhibitors; structure-activity relationship.