One-pot multi-component synthesis and biological evaluation of spirooxindole carbamate derivatives of major flavonoids isolated from oroxylum indicum: Identification of potent anti-proliferative leads active against hepatocellular carcinoma.

A series of 30 novel spirooxindole carbamate derivatives of flavonoids (chrysin, oroxylin A and baicailein) were synthesized in a one-pot three-component reaction via a sequence of Knoevenagel condensation, cyclization reactions in a highly efficient and selective manner. The structures of all the derivatives (6a-6 l, 7a-7 l and 8a-8f) were characterized by interpretation of NMR (H NMR, C NMR) and mass spectral data. The structures of the compounds 6d and 7f were further confirmed by single-crystal X-ray diffraction for the first time. The anti-proliferative potential of these derivatives was assessed against various cancer cells such as A549 (lung), MDA-MB-231 (breast), HeLa (cervical), HepG2 (liver), and HEK 293 (normal kidney) cell lines using the Sulforhodamine-B assay. Among all tested compounds, 6d exhibited the most potent anti-proliferative activity, with an IC₅₀ of 2.50 ± 0.25 μM against the HepG2 cell line. Compound 6d significantly enhanced early apoptosis in comparison to control cells and markedly (p < 0.001) reduced the population of CD133 cancer stem-like cells, which are critical mediators of metastasis and drug resistance. Molecular docking studies revealed a strong interactions between compound 6d and critical residues of CD133, including Phe787, Val794, Phe62, Lys791, and Leu5, with a binding affinity of -10.2 kcal/mol. Furthermore, compound 6d treatment reduced sphere formation in a dose-dependent manner in HepG2 cells. In vivo studies using a xenograft model demonstrated that compound 6d significantly suppressed tumor growth and reduced tumor weight, indicating its potential as a promising therapeutic candidate for hepatocellular carcinoma.