Design, synthesis and biological evaluation of (±)-kusunokinin derivatives as potent anticancer agents

Kusunokinin and its derivatives, which possess a dibenzylbutyrolactone core, have demonstrated notable anticancer activity, particularly against breast, ovarian, and cholangiocarcinoma cell lines. In this study, a series of (±)-kusunokinin derivatives were rationally designed based on previously reported bioactivity data to investigate structure–activity relationships (SAR) at the R, R′, and R″ positions of the aromatic rings. Structural modifications included the introduction of hydrogen bond donors, hydrogen bond acceptors, aromatic π-systems, and lipophilic groups. The influence of a methoxy substituent at the R″ position was also systematically evaluated. The derivatives were synthesized via a previously established route, utilizing (±)-intermediates 9 and 12 as the key building block. Hydroxyl protection and deprotection strategies were introduced, enabling selective functionalization of specific hydroxyl groups on the aromatic rings. Four compounds ( 13 , 16 , 18 , and 33 ) exhibited cytotoxic activity against cholangiocarcinoma, triple-negative breast cancer, and ER-positive ductal carcinoma cell lines, while demonstrating lower cytotoxicity toward normal cells. Derivatives featuring an alkoxy or phenyl substituent at the R position and a hydroxyl group at R′ demonstrated enhanced cytotoxic activity. In contrast, the contribution of the methoxy group at R″ remains unclear and warrants further investigation. A discrepancy between the in silico screening results and experimental findings was also observed. These findings support the continued development of dibenzylbutyrolactone-based scaffolds as promising anticancer agents via targeting cancer-associated kinase. • The newly developed synthetic platform employed protection and deprotection of the hydroxyl group as a key strategy to prepare eighteen (±)-kusunokinin derivatives, while the remaining four derivatives were synthesized via the original pathway. • Twenty-two (±)-kusunokinin derivatives were systematically designed by modifying the functional groups and altering the position of the hydroxyl group at the R and R′ sites. • (±)-Kusunokinin derivatives bearing a hydroxyl group at the R′ position and an alkoxy chain or a phenyl ring at the R position exhibited higher cytotoxicity against cancer cells than the other derivatives. • Four compounds ( 13 , 16 , 18 , and 33 ) demonstrated potent and selective cytotoxicity against cholangiocarcinoma, triple-negative breast cancer, and ER-positive ductal carcinoma cell lines, while sparing normal cells—underscoring their potential as promising anticancer candidates. • A new approach focusing on kinase targeting in cancer pathways, combined with improved water solubility, could be the next direction for kusunokinin modification, rather than focusing solely on CSF1R and/or AKR1B1.