Highly efficacious and tolerable agents for the treatment of glioblastoma (GBM), the most common and aggressive primary brain tumor, are urgently needed. Herein, we reveal the design, synthesis and biological evaluation of several piperazine based benzamide derivatives, which are based on the non-classical isostere principle and combination principle for GBM therapy. After structure-activity relationship (SAR) study, compound L19 was demonstrated as the most promising compound with IC₅₀ values of 0.15 μM, 0.29 μM, 1.25 μM against GBM C6, U87-MG, U251 cells, respectively. Moreover, compound L19 could inhibit the proliferation, migration and invasion, as well as induce apoptosis and cell cycle arrest of GBM cell lines in vitro. From mechanism perspective, compound L19 could regulate the cell cycle-related proteins and influence the p16^INK4a-CDK4/6-pRb pathway by western blotting experiment. What is worth mentioning is that compound L19 could penetrate the blood-brain barrier (BBB) with an exceptional brain-to-plasma ratio of 1.07 in vivo. Besides, the superior anti-glioblastoma potency in vivo of compound L19 was identified on U87-MG-xenograft model without any apparent host toxicity. Overall, the potential of compound L19 warrants further pre-clinical investigation for GBM therapy.

迫切需要用于治疗最常见和最具侵袭性的原发性脑肿瘤胶质母细胞瘤 (GBM) 的高效和耐受药物。在此，我们揭示了几种基于哌嗪的苯甲酰胺衍生物的设计、合成和生物学评价，这些衍生物基于 GBM 治疗的非经典等排原理和组合原理。经过构效关系 (SAR) 研究，化合物L19被证明是最有希望的化合物，其对 GBM C6、U87-MG、U251 细胞的 IC ₅₀值分别为 0.15 μM、0.29 μM、1.25 μM。此外，化合物L19在体外可抑制GBM细胞系的增殖、迁移和侵袭，诱导细胞凋亡和细胞周期停滞。. 从机制上看，通过蛋白质印迹实验，化合物L19可以调节细胞周期相关蛋白并影响p16 ^{INK4a -CDK4/6-pRb通路。}值得一提的是，化合物L19可以穿透血脑屏障 (BBB)，在体内具有 1.07 的出色脑浆比。此外，在U87-MG-异种移植模型上鉴定出化合物L19在体内具有优异的抗胶质母细胞瘤效力，没有任何明显的宿主毒性。总体而言，化合物L19的潜力值得进一步进行 GBM 治疗的临床前研究。