Tubulin polymerization-IN-4 is an effective inhibitor of tubulin polymerization (IC50=4.6 μM). Tubulin polymerization-IN-4 can destroy tubulin polymerization, block cell cycle at G2/M phase, induce apoptosis, and inhibit cell cloning and migration. Tubulin polymerization-IN-4 can also damage the vascular system.

Tubulin polymerization-IN-4 (compound 9j) exhibits significant antitumor activity by inhibiting cancer cell activity and tubulin polymerization. It demonstrates sub-micromolar inhibitory effects against HeLa, SiHa, and MS751 cell lines (0-1 μM, 48 hours), with notable IC50 values. At higher concentrations (3, 6, and 12.5 μM, 0-20 minutes), it inhibits tubulin polymerization in a dose-dependent manner (39%, 54%, and 77% inhibition, respectively). The compound also interferes with EBI-β-tubulin adduct formation and disrupts vascular tube formation in HUVEC cells, indicating potential anti-angiogenic effects. At low micromolar concentrations (0.1-0.4 μM, 24 hours), it induces G2/M phase arrest, triggers apoptosis, and inhibits migration and colony formation in HeLa cells. A cytotoxicity assay in HK-2 cells reveals a favorable renal safety profile (IC50 of 188 ± 16 μM). These findings highlight Tubulin polymerization-IN-4 as a promising therapeutic agent with a well-defined mechanism of action and acceptable safety profile.

Tubulin polymerization-IN-4, administered intraperitoneally (IP) at doses ranging from 100 to 1000 mg/kg, exhibits very low toxicity, with its lethal dose 50 (LD50) exceeding 1000 mg/kg [1]. When given daily for 21 days at doses of 30 and 60 mg/kg, it effectively inhibits tumor growth, achieving tumor growth inhibition (TGI) rates of 35% and 58%, respectively, at these doses [1]. Furthermore, a single IP administration of Tubulin polymerization-IN-4 at 30 mg/kg demonstrates moderate pharmacokinetic properties, with detailed parameters in ICR mice including a half-life (T 1/2) of 1.56 ± 0.28 hours, a time to reach maximum concentration (T max) of 0.25 hours, a maximum concentration (C max) of 6215 ± 308 μg/L, an area under the curve from 0 to last measurable concentration (AUC 0-t) of 5609 ± 347 μg/L·h, an area under the curve from 0 to infinity (AUC 0-∞) of 5940 ± 347 μg/L·h, a volume of distribution (V Z /F) of 11.35 ± 1.29 L/kg, a clearance rate (CL Z /F) of 5.05 ± 0.91 L/h/kg, and a mean residence time (MRT) of 1.77 ± 0.43 hours [1].