Key Laboratory of Neuropsychiatric Drug Research of Zhejiang Province, Institute of Materia Medica, Hangzhou Medical College, Hangzhou, Zhejiang 310013, China.
Key Laboratory of Neuropsychiatric Drug Research of Zhejiang Province, Institute of Materia Medica, Hangzhou Medical College, Hangzhou, Zhejiang 310013, China.
Bioorg Med Chem. 2021 Jun 1;39:116166. doi: 10.1016/j.bmc.2021.116166. Epub 2021 Apr 21.
Constitutive activation of Hedgehog (Hh) pathway is intimately related with the occurrence and development of several malignancies, such as medulloblastoma (MB) and other tumors. Therefore, small molecular inhibitors of Hh pathway are urgently needed. In this study, three new steroidal alkaloids, ⊿ (20R, 24R) 23-oxo-24-methylsolacongetidine, ⊿ (20S, 24R) 23-oxo-24-methylsolacongetidine and veralinine 3-O-α-l-rhamnopyranosyl-(1 → 2)-β-D-glucopyranoside, together with six known alkaloids, 20-epi-verazine, verazine, protoverine 15-(l)-2'-methylbutyrate, jervine, veramarine and β1-chaconine, were isolated and determined from Veratrum grandiflorum Loes. The dual-luciferase bioassay indicated that all compounds exhibited significant inhibitions of Hh pathway with IC values of 0.72-14.31 μM against Shh-LIGHT 2 cells. To determine whether these Hh pathway inhibitors act with the Smoothened (Smo) protein, which is an important oncoprotein and target for this pathway, BODIPY-cyclopamine (BC) competitive binding assay was preferentially performed. Compared with BC alone, all compounds obviously reduced the fluorescence intensities of BC binding with Smo in Smo-overexpression HEK293T cells through fluorescence microscope and flow cytometer. By directly interacting with Smo, it revealed that they were actually novel natural Smo inhibitors. Then, their anti-tumor effects were investigated against the human MB cell line DAOY, which is a typical pediatric brain tumor cells line with highly expressed Hh pathway. Interestingly, most of compounds had slight proliferation inhibitions on DAOY cells after treatment for 24 h same as vismodegib, while β1-chaconine showed the strongest inhibitory effect on the growth of DAOY with IC value of 5.35 μM. In conclusion, our studies valuably provide several novel natural Smo inhibitors for potential targeting treatment of Hh-dependent tumors.
Hedgehog (Hh) 信号通路的组成性激活与几种恶性肿瘤的发生和发展密切相关，如髓母细胞瘤 (MB) 和其他肿瘤。因此，迫切需要 Hh 通路的小分子抑制剂。在这项研究中，从藜芦属植物中分离得到了三种新的甾体生物碱，⊿(20R, 24R)23-氧代-24-甲基索尔康替丁、⊿(20S, 24R)23-氧代-24-甲基索尔康替丁和 veralinine 3-O-α-l-鼠李吡喃糖基-(1→2)-β-D-葡萄糖苷，以及 6 种已知的生物碱，20-表-verazine、verazine、protoverine 15-(l)-2'-甲基丁酸酯、jervine、veramarine 和 β1-chaconine。双荧光素酶生物测定表明，所有化合物对 Shh-LIGHT 2 细胞均显示出显著的 Hh 通路抑制活性，IC 值为 0.72-14.31 μM。为了确定这些 Hh 通路抑制剂是否与 Smoothened (Smo) 蛋白作用，Smo 蛋白是该通路的重要癌蛋白和靶标，因此优先进行了 BODIPY-cyclopamine (BC) 竞争性结合测定。与单独的 BC 相比，所有化合物在 Smo 过表达的 HEK293T 细胞中通过荧光显微镜和流式细胞仪明显降低了 BC 与 Smo 结合的荧光强度。通过与 Smo 直接相互作用，表明它们实际上是新型天然 Smo 抑制剂。然后，研究了它们对人 MB 细胞系 DAOY 的抗肿瘤作用，DAOY 是一种典型的小儿脑肿瘤细胞系，Hh 通路高度表达。有趣的是，与 vismodegib 一样，大多数化合物在处理 24 小时后对 DAOY 细胞的增殖抑制作用轻微，而 β1-chaconine 对 DAOY 的生长抑制作用最强，IC 值为 5.35 μM。总之，我们的研究为潜在的针对 Hh 依赖性肿瘤的治疗提供了有价值的几种新型天然 Smo 抑制剂。
