从淡竹叶中分离得到的倍半萜对人乳腺癌细胞迁移的体外抑制作用及其LIMK1 下调机制

In vitro inhibitory properties of sesquiterpenes from Chloranthus serratus on cell motility via down-regulation of LIMK1 activation in human breast cancer.

机构信息

Department of Pharmacology, School of Pharmacy, Jiangxi University of Traditional Chinese Medicine, Nanchang 330004, China.

出版信息

Phytomedicine. 2018 Oct 1;49:23-31. doi: 10.1016/j.phymed.2018.06.009. Epub 2018 Jun 13.

DOI:10.1016/j.phymed.2018.06.009

PMID:30217259

Abstract

BACKGROUND

LIM kinase 1 plays an important role in tumor cell invasion and metastasis by regulating architecture of actin cytoskeleton, and inhibiting activity of this kinase may be a promising strategy to prevent cancer cells from distant spread. In our previous studies, we found several extracts from the medical herbs in genus Chloranthus to exhibit anti-metastatic effects.

PURPOSE

The aim of this study is to find LIMK1 inhibitors from Chloranthus serratus, a medical herb from genus Chloranthus and to evaluate their effects on cell motility.

METHODS

Three sesquiterpenes, chloranthalactone E (compound 1), serralactone A (compound 2, SERA is used in the further testing), and 8β, 9α-dihydroxylindan-4(5), 7(11)-dien-8α, 12-olide (compound 3) were isolated from Chloranthus serratus, and the anti-LIMK1 activities of these compounds were investigated by kinase-Glo luminescent kinase assay. Then, the anti-LIMK1 properties of SERA were verified by kinase-Glo luminescent kinase assay and western blot assay. The effects of SERA on F-actin polymerization and cell migration were investigated by Phalloidin dying, AP 48 chamber system and ORIS™ cell migration assay. Furthermore, the inhibitory effects of SERA on LIMK1 were confirmed by overexpression of LIMK1 and small interfering RNA (siRNA) mediated gene silencing.

RESULTS

we reported here that among the three sesquiterpenes, SERA showed significantly inhibition on LIMK1 activity, and the IC values on MDA-MB-231 and MDA-MB-468 cells were 3.14 μM and 4.64 μM, respectively. Furthermore, it was also found that SERA significantly suppressed LIMK1 and cofilin1 phosphorylation, F-actin polymerization and also cell migration. Data from LIMK1 overexpression and RNA interfering assay confirmed that the inhibitory effects of SERA on LIMK1 was antagonized and enhanced by the overexpression and knockdown of LIMK1.

CONCLUSION

collectively, it was concluded that SERA exhibited significant inhibitory effects on breast cancer cells migration, and these effects of this sesquiterpene are due to its properties reducing the activation of LIM kinase 1.

摘要

背景

LIM 激酶 1 通过调节肌动蛋白细胞骨架的结构在肿瘤细胞侵袭和转移中发挥重要作用，抑制该激酶的活性可能是防止癌细胞远处扩散的一种有前途的策略。在我们之前的研究中，我们发现几种来自金粟兰属药用植物的提取物具有抗转移作用。

目的

本研究的目的是从金粟兰属药用植物金粟兰中寻找 LIMK1 抑制剂，并评价其对细胞迁移的影响。

方法

从金粟兰中分离得到三种倍半萜类化合物，即金粟兰内酯 E（化合物 1）、金粟兰内酯 A（化合物 2，SERA 用于进一步测试）和 8β,9α-二羟基-4(5),7(11)-二烯-8α,12-内酯（化合物 3），通过激酶-Glo 发光激酶测定法研究这些化合物的抗 LIMK1 活性。然后，通过激酶-Glo 发光激酶测定法和 Western blot 测定法验证 SERA 的抗 LIMK1 特性。通过鬼笔环肽染色、AP48 室系统和 ORIS™细胞迁移测定法研究 SERA 对 F-肌动蛋白聚合和细胞迁移的影响。此外，通过 LIMK1 的过表达和小干扰 RNA（siRNA）介导的基因沉默，证实 SERA 对 LIMK1 的抑制作用。

结果

我们在这里报道，在这三种倍半萜中，SERA 对 LIMK1 活性表现出显著的抑制作用，对 MDA-MB-231 和 MDA-MB-468 细胞的 IC 值分别为 3.14 μM 和 4.64 μM。此外，还发现 SERA 显著抑制 LIMK1 和丝切蛋白 1 磷酸化、F-肌动蛋白聚合和细胞迁移。LIMK1 过表达和 RNA 干扰测定的数据证实，SERA 对 LIMK1 的抑制作用被 LIMK1 的过表达和敲低拮抗和增强。