Design, synthesis and biological study of hybrid drug candidates of nitric oxide releasing cucurbitacin-inspired estrone analogs for treatment of hepatocellular carcinoma.

Development of hybrid drug candidates is well known strategy for designing antitumor agents. Herein, a novel class of nitric oxide donating cucurbitacin inspired estrone analogs (NO-CIEAs) were designed and synthesized as multitarget agents. Synthesized analogs were initially evaluated for their anti-hepatocellular carcinoma activities. Among the tested analogs, NO-CIEAs 17 and 20a exhibited more potent activity against HepG2 cells (IC = 4.69 and 12.5 µM, respectively) than the reference drug Erlotinib (IC = 25 µM). Interestingly, NO-CIEA 17 exerted also a high potent activity against Erlotinib-resistant HepG2 cell line (HepG2-R) (IC = 8.21 µM) giving insight about its importance in drug resistance therapy. Intracellular measurements of NO revealed that NO-CIEAs 17 and 20a showed a significant increase in NO production in tumor cells after 1 h of incubation comparable to the reference prodrug JS-K. Flow cytometric analysis showed that both NO-CIEAs 17 and 20a mainly arrested the HepG2 cells in the G0/G1 phase. Also, In-Cell Based ELISA screening showed that NO-CIEA 17 resulted in a potential inhibitory activity towards the EGFR and MAPK (25% and 29% inhibition compared to untreated control cells, respectively). This data suggests the binding ability of NO-CIEA 17 to the EGFR and ERK to be well correlated along with the docking and cellular studies. Also, treatment of HepG2-R cells with NO-CIEA 17 showed a potential reduction of MRP2 expression in a dose dependent manner providing a significant impact on the chemotherapeutic resistance. Overall, the current study provides a potential new approach for the discovery of a novel antitumor agent against HCC.