Development of a Novel Indirubin Derivative with Enhanced Anticancer Properties: Synthesis, In Vitro, and In Vivo Evaluation

Abstract

Abstract The indirubin (2) derivative undergoes condensation with hydroxylamine hydrochloride, resulting in the formation of a novel indirubin derivative (3) that incorporates both the oxime group and the piperidine heterocycle. This compound is subsequently treated with hydrochloric acid in ethanol, leading to the formation of the hydrochloride salt (4), which exhibits excellent solubility in water. The chemical structures of compounds (3) and (4) were determined using 1D and 2D nuclear magnetic resonance (NMR) spectroscopy, as well as high-resolution mass spectrometry with electrospray ionization (ESI). To evaluate their potential as anticancer agents, in vitro assays were conducted using three human cancer cell lines (A549, Hep-G2, SW480) and one murine cancer cell line (B16F10). These compounds, along with indirubin (1) and Bortezomib (BTZ), were tested. The results revealed that compounds (3) and (4) demonstrated significant antitumor activity against all four cancer cell lines, with concentrations ranging from 0.363 to 12.990 µM. Importantly, their activity surpassed that of indirubin (1) and BTZ. Furthermore, an in vivo anticancer activity assay was performed on compound (4) using the B16F10 cell line. After 18 days, the tumor size in the group treated with compound (4) was approximately eight times smaller than that of the control group, and nearly three times smaller than the group treated with BTZ.