Inhibitory Effect of Liuwei Dihuangwan on Triple-negative Breast Cancer Through MAPKKK1 and KLF4 / 中国实验方剂学杂志

ABSTRACT

ObjectiveTo study the underlying mechanism of Liuwei Dihuangwan in inhibiting triple-negative breast cancer through mitogen-activated protein kinase kinase kinase 1 （MAPKKK1） and Krüppel-like factor 4 （KLF4）. MethodFour hundreds SPF female Kunming mice aged 11.5 months were palpated once every 3 days. The model mice of spontaneous tumors were randomly divided into a model group （normal saline）， a paclitaxel group （0.025 g·kg-1·d-1， ip， 21 days）， and high-， medium- and low-dose Liuwei Dihuangwan groups （7.2， 3.6， 1.8 g·kg-1·d-1， ig）. Tumor tissues were separated until the moribund stage. The tumor volume and weight were measured， and the tumor inhibition rate and the survival time of the tumor mice were calculated （after 6 months， tumor-free mice were assigned into the normal group）. SPF SD rats were selected to prepare serum samples containing Liuwei Dihuangwan of different concentrations for cell culture， and MAPKKK1 in MDA-MB-231 cells was silenced. The protein expression of MAPKKK1 and KLF4 was detected by immunofluorescence and Western blot. ResultThe in vivo experimental results showed that compared with the conditions of the normal group， the protein expression of MAPKKK1 and KLF4 in tumor tissues of the model group dropped （P<0.01）. Compared with the model group， all medication groups showed reduced tumor volume and weight （P<0.05， P<0.01）， increased tumor inhibition rate， prolonged survival time of tumor mice （P<0.05）， and increased protein expression of MAPKKK1 （P<0.01）. Additionally， the paclitaxel group and the high-dose Liuwei Dihuangwan group exhibited increased protein expression of KLF4 （P<0.01）. The in vitro experiments showed that compared with the conditions of the normal group， the fluorescence intensities of MAPKKK1 and KLF4 in MDA-MB-231 cells in all medication groups were potentiated， and the protein expression of MAPKKK1 in the paclitaxel group and the high- and medium-dose Liuwei Dihuangwan groups， and the protein expression of KLF4 in the paclitaxel group and high-dose Liuwei Dihuangwan group increased （P<0.01）. After silencing of MAPKKK1， compared with the conditions of the negative plasmid group （unsilenced MAPKKK1）， the fluorescence intensities of MAPKKK1 and KLF4 and the protein expression decreased in the RNAi-27 positive plasmid group （silenced MAPKKK1） （P<0.05， P<0.01）. Compared with the RNAi-27 positive plasmid group， all medication groups had enhanced fluorescence intensities of MAPKKK1 and KLF4 and protein expression to different degrees （P<0.05， P<0.01）. ConclusionLiuwei Dihuangwan can inhibit the growth of triple-negative breast cancer， and the underlying molecular mechanism is related to the up-regulation of MAPKKK1 and activation of KLF4 expression.