As one of the classical traditional Chinese medicine (TCM) prescriptions in treating gynecological tumors, Guizhi Fuling Decoction (GFD) has been used to treat breast cancer (BRCA). Nonetheless, the potential molecular mechanism remains unclear so far. Therefore, systems pharmacology was used in combination with high throughput sequencing-based high throughput screening (HTS²) assay and bioinformatic technologies in this study to investigate the molecular mechanisms of GFD in treating BRCA. By computationally analyzing 76 active ingredients in GFD, 38 potential therapeutic targets were predicted and significantly enriched in the “pathways in cancer”. Meanwhile, experimental analysis was carried out to examine changes in the expression levels of 308 genes involved in the “pathways in cancer” in BRCA cells treated by five herbs of GFD utilizing HTS² platform, and 5 key therapeutic targets, including HRAS, EGFR, PTK2, SOS1, and ITGB1, were identified. The binding mode of active compounds to these five targets was analyzed by molecular docking and molecular dynamics simulation. It was found after integrating the computational and experimental data that, GFD possessed the anti-proliferation, pro-apoptosis, and anti-angiogenesis activities mainly through regulating the PI3K and the MAPK signaling pathways to inhibit BRCA. Besides, consistent with the TCM theory about the synergy of Cinnamomi Ramulus (Guizhi) by Cortex Moutan (Mudanpi) in GFD, both of these two herbs acted on the same targets and pathways. Taken together, the combined application of computational systems pharmacology techniques and experimental HTS² platform provides a practical research strategy to investigate the functional and biological mechanisms of the complicated TCM prescriptions.

作为治疗妇科肿瘤的经典中药（TCM）处方之一，桂枝Fu灵汤（GFD）已被用于治疗乳腺癌（BRCA）。尽管如此，到目前为止，潜在的分子机制仍不清楚。因此，系统药理学与基于高通量测序的高通量筛选（HTS ²）分析和生物信息技术，以研究GFD治疗BRCA的分子机制。通过计算分析GFD中的76种有效成分，可以预测38种潜在的治疗靶点，并且这些靶点在“癌症的途径”中显着丰富。同时，进行了实验分析，以检查五种GFD草药利用HTS ²处理的BRCA细胞中“癌症途径”中308个基因的表达水平的变化。平台，并确定了5个关键治疗靶点，包括HRAS，EGFR，PTK2，SOS1和ITGB1。通过分子对接和分子动力学模拟分析了活性化合物与这五个靶标的结合方式。综合计算和实验数据发现，GFD主要通过调节PI3K和MAPK信号通路抑制BRCA而具有抗增殖，促凋亡和抗血管生成的作用。此外，有关的协同中医理论是一致肉桂桂枝（桂枝）由牡丹皮（牡丹皮）在GFD，既作用于同一个目标和途径这两种草药。综合起来，计算系统药理学技术与实验性HTS ²的组合应用 该平台为研究复杂中药处方的功能和生物学机制提供了实用的研究策略。