甲状腺杨河汤（JWYHD）是临床上用于治疗自身免疫性疾病的广泛应用的传统中药方剂。许多研究表明，JWYHD在细胞和动物模型中具有抗肿瘤活性。然而，JWYHD的抗乳腺癌作用及作用机制仍然未知。本研究旨在确定JWYHD的抗乳腺癌效应并揭示其在体内、体外和体外的作用机制。采用原位移植的乳腺癌小鼠模型和炎症斑马鱼模型观察JWYHD的抗肿瘤作用和免疫细胞调节作用。此外，通过RAW 264.7细胞的表达评估JWYHD的抗炎作用。使用UPLC-MS/MS获得JWYHD活性成分，并由网络药理学筛选潜在靶点。最后，通过Western blot，实时PCR（RT-PCR），免疫组织化学（IHC）染色和酶联免疫吸附实验（ELISA）评估计算机预测的治疗靶点和信号通路，以探索JWYHD对乳腺癌的治疗机制。JWYHD在原位移植的乳腺癌小鼠模型中呈剂量依赖性地显著降低了肿瘤生长。流式细胞术和IHC结果表明，JWYHD降低了M2巨噬细胞和Treg的表达，同时增加了M1巨噬细胞。同时，ELISA和Western blot结果显示JWYHD组的肿瘤组织中IL-1β，IL-6，TNFα，PTGS2和VEGFα的表达减少。结果在LPS诱导的RAW264.7细胞和斑马鱼炎性模型中得到验证。TUNEL分析和IHC结果表明JWYHD能够显著诱导细胞凋亡。使用UPLC-MS/MS和网络药理学确定了JWYHD的72个主要化合物。发现JWYHD与TNFα，PTGS2，EGFR，STAT3，VEGFα具有显著的结合亲和力，且JWYHD可抑制它们的表达。Western blot和IHC分析证实JWYHD通过调节JAK2 / STAT3信号通路在抗肿瘤和免疫调节中发挥重要作用。JWYHD主要通过抑制炎症、激活免疫反应和通过JAK2 / STAT3信号通路诱导细胞凋亡来产生显著的抗肿瘤效应。我们的发现为JWYHD在治疗乳腺癌中的临床应用提供了强有力的药理学证据。版权所有©2023 Elsevier BV.

Jiawei Yanghe Decoction (JWYHD) is a widely used traditional Chinese medicine prescription in the clinical setting for the treatment of autoimmune diseases. Many studies showed that JWYHD has anti-tumor activities in cell and animal models. However, the anti-breast cancer effects of JWYHD and the underlying mechanisms of action remain unknown.This study aimed to determine the anti-breast cancer effect and reveal the underlying mechanisms of action in vivo, in vitro and in silico.Orthotopic xenograft breast cancer mouse model and inflammatory zebrafish model were used to observe the anti-tumor effect and immune cell regulation of JWYHD. Moreover, the anti-inflammatory effect of JWYHD were evaluated by the expression of RAW 264.7 cells. JWYHD active ingredients were obtained by UPLC-MS/MS and potential targets were screened by network pharmacology. At last, the therapeutic targets and signaling pathways predicted by computer were assessed by western blot, real-time PCR (RT-PCR), immunohistochemistry (IHC) staining, and Enzyme-linked immunosorbent assays (ELISA) to explore the the therapeutic mechanism of JWYHD against breast cancer.JWYHD significantly decreased the tumor growth in a dose-dependent manner in the orthotopic xenograft breast cancer mouse model. Flow cytometry and IHC results indicated that JWYHD decreased the expressions of M2 macrophages and Treg while increasing M1 macrophages. Meanwhile, ELISA and western blot results showed a decrease in IL-1β, IL-6, TNFα, PTGS2 and VEGFα in tumor tissue of JWYHD groups. The results were also verified in LPS-induced RAW264.7 cells and zebrafish inflammatory models. TUNEL assay and IHC results showed that JWYHD significantly induced apoptosis. Seventy-two major compounds in JWYHD were identified by UPLC-MS/MS and Network pharmacology. It was found that the significant binding affinity of JWYHD to TNFα, PTGS2, EGFR, STAT3, VEGFα and their expression was inhibited by JWYHD. Western blot and IHC analysis confirmed that JWYHD plays an important role in anti-tumor and immune regulation by regulating JAK2/STAT3 signaling pathway.JWYHD exerts a significant anti-tumor effect mainly by inhibiting inflammation, activating immune responses and inducing apoptosis via the JAK2/STAT3 signaling pathway. Our findings provide strong pharmacological evidence for the clinical application of JWYHD in the management of breast cancer.Copyright © 2023. Published by Elsevier B.V.