Discovery of New Quinazolinone and Benzimidazole Analogs as Tubulin Polymerization Inhibitors with Potent Anticancer Activities

<b>Background/Objectives:</b> Cancer persists as a leading concern in the current medical field, and current therapies are limited by toxicity, cost, and resistance. Targeted inhibition of tubulin polymerization is considered as a promising therapeutic strategy for cancer treatment. <b>Methods</b>: Thirty-one new tubulin polymerization inhibitors were designed via molecular hybridization techniques, and BLI technology was employed to quantitatively investigate their interactions with tubulin. Antiproliferative activities against MCF-7, MDA-MB-231, A549, and HeLa cell lines was evaluated using the CCK8 assay. Apoptosis induction and cell cycle arrest were analyzed by flow cytometry. The anti-tumor activity of compound <b>B6</b> was validated in a mouse melanoma tumor model. <b>Results</b>: Compounds exhibited varying degrees of antiproliferative activity against four tumor cell lines. Among them, compound <b>B6</b> was the most promising candidate and displayed strong broad-spectrum anticancer activity with an average IC₅₀ value of 2 μM. The mechanism studies revealed that compound <b>B6</b> inhibited tubulin polymerization in vitro, disrupted cell microtubule networks, and arrested the cell cycle at G2/M phase. Furthermore, <b>B6</b> displayed significant in vivo antitumor efficacy in a melanoma tumor model with tumor growth inhibition rates of 70.21% (50 mg/kg). <b>Conclusions</b>: This work shows that <b>B6</b> is a promising lead compound deserving further investigation as a potential anticancer agent.