Department of Chemistry and Technologies of Drugs, Sapienza University of Rome, P.le A. Moro 5, 00185, Rome, Italy.
Department of Experimental Medicine, Sapienza University of Rome, P.le A. Moro 5, 00185, Rome, Italy.
Clin Epigenetics. 2019 Dec 2;11(1):173. doi: 10.1186/s13148-019-0763-5.
Glioblastoma (GBM) is the most lethal and aggressive malignant primary brain tumor in adults. After surgical resection of the tumor, the patient typically should be subjected to chemotherapy (temozolomide, TMZ) and concomitant radiotherapy. Since the TMZ treatment does not lead to complete remission and often develops resistance, the identification of efficacious therapeutics is strongly to pursue. Among the epigenetic players, the H3K27 methyltransferase (MT) EZH2 (enhancer of zeste homologue 2) has been found overexpressed or mutated in several human cancers including gliomas, and its overexpression is associated with poor outcome in GBM. Two EZH2 inhibitors (EZH2i), UNC1999 and GSK343, suppressed GBM growth in vitro and in vivo indicating that EZH2i can be potential drugs against GBM.
Two new EZH2i, MC4040 and MC4041, were designed, prepared, and tested by us to determine their effects in primary GBM cell cultures. MC4040 and MC4041 displayed single-digit micromolar inhibition of EZH2, 10-fold less potency against EZH1, and no activity towards other MTs. In primary GBM cells as well as in U-87 GBM cells, the two compounds reduced H3K27me3 levels, and dose- and time-dependently impaired GBM cell viability without inducing apoptosis and arresting the cell cycle in the G0/G1 phase, with increased p21 and p27 levels. In combination with TMZ, MC4040 and MC4041 displayed stronger, but not additive, effects on cell viability. The potent clinical candidate as EZH2i tazemetostat, alone or in combination with TMZ, exhibited a similar potency of inhibition of GBM cell growth when compared to MC4040 and MC4041. At the molecular level, MC4040 and MC4041 reduced the VEGFR1/VEGF expression, reversed the epithelial-mesenchymal transition (EMT), and hampered cell migration and invasion attenuating the cancer malignant phenotype. Treatment of GBM cells with MC4040 and MC4041 also impaired the GBM pro-inflammatory phenotype, with a significant decrease of TGF-β, TNF-α, and IL-6, joined to an increase of the anti-inflammatory cytokine IL-10.
The two novel EZH2i MC4040 and MC4041 impaired primary GBM cell viability, showing even stronger effects in combination with TMZ. They also weakened the aggressive malignant phenotype by reducing angiogenesis, EMT, cell migration/invasion and inflammation, thus they may be considered potential candidates against GBM also for combination therapies.
胶质母细胞瘤（GBM）是成人中最致命和侵袭性的恶性原发性脑肿瘤。在肿瘤切除术后，患者通常应接受化疗（替莫唑胺，TMZ）和同步放疗。由于 TMZ 治疗不能导致完全缓解，并且常常产生耐药性，因此强烈需要寻找有效的治疗方法。在表观遗传因子中，H3K27 甲基转移酶（MT）EZH2（增强子的同系物 2）在包括神经胶质瘤在内的几种人类癌症中发现过度表达或突变，其过度表达与 GBM 的不良预后相关。两种 EZH2 抑制剂（EZH2i）UNC1999 和 GSK343 在体外和体内均抑制 GBM 生长，表明 EZH2i 可成为治疗 GBM 的潜在药物。
我们设计、制备和测试了两种新型 EZH2i，即 MC4040 和 MC4041，以确定它们在原发性 GBM 细胞培养物中的作用。MC4040 和 MC4041 对 EZH2 的抑制作用呈个位数微摩尔，对 EZH1 的抑制作用强 10 倍，对其他 MT 无活性。在原发性 GBM 细胞和 U-87 GBM 细胞中，两种化合物均降低了 H3K27me3 水平，并剂量和时间依赖性地损害了 GBM 细胞活力，而不会诱导细胞凋亡和将细胞周期阻滞在 G0/G1 期，同时增加了 p21 和 p27 水平。与 TMZ 联合使用时，MC4040 和 MC4041 对细胞活力的抑制作用更强，但并非相加。作为 EZH2i 的有效临床候选药物 tazemetostat 与 MC4040 和 MC4041 相比，单独或与 TMZ 联合使用时，对 GBM 细胞生长的抑制作用相似。在分子水平上，MC4040 和 MC4041 降低了 VEGFR1/VEGF 的表达，逆转了上皮-间充质转化（EMT），并阻碍了细胞迁移和侵袭，从而削弱了癌症的恶性表型。用 MC4040 和 MC4041 处理 GBM 细胞也削弱了 GBM 的促炎表型，使 TGF-β、TNF-α 和 IL-6 显著减少，同时增加抗炎细胞因子 IL-10。
两种新型 EZH2i MC4040 和 MC4041 损害了原发性 GBM 细胞活力，与 TMZ 联合使用时效果更强。它们还通过减少血管生成、EMT、细胞迁移/侵袭和炎症来减弱侵袭性恶性表型，因此它们可能被认为是治疗 GBM 的潜在候选药物，也可用于联合治疗。
