【目的】 基于网络药理学、分子对接及细胞实验验证探讨二至丸治疗多发性骨髓瘤（MM）的潜在作用机制。【方法】 应用 网络药理学、分子对接筛选药物-疾病-靶标-通路关键指标。培养多发性骨髓瘤细胞株RPMI-8226，给予二至丸干预，采用 膜联蛋白V（Annexin V）-异硫氰酸荧光素（FITC）染色法检测细胞凋亡，Transwell实验测定细胞侵袭能力，实时定量聚合酶链 反应（qRT-PCR）法检测蛋白激酶 B（Akt）、细胞周期蛋白 D1（CCND1）、糖原合酶激酶 3β（GSK-3β）、c-Myc mRNA 表达， Western Blot 法检测细胞 Akt、磷酸化蛋白激酶 B（p-Akt）、CCND1、c-Myc、GSK-3β、磷酸化糖原合酶激酶 3β（p-GSK-3β） 蛋白表达。【结果】 获得二至丸14个有效成分。二至丸治疗MM涉及靶点142个，通路富集分析结果显示关键靶点可能主要集 中在癌症通路、脂质与动脉粥样硬化等。分子对接结果显示，木樨草素和槲皮素与GSK-3β有较好的结合活性及稳定性。进 一步细胞实验验证发现，与空白组比较，二至丸低、中、高剂量组细胞凋亡率显著升高，细胞侵袭数减少，GSK-3β 的 mRNA和蛋白水平显著升高，CCND1、Akt和c-Myc的mRNA和蛋白水平显著降低，呈剂量依赖性，其中，中、高剂量组差 异均有统计学意义（P＜0.05或P＜0.01）。【结论】 二至丸对MM的治疗作用可能是通过木犀草素和槲皮素等关键活性成分，促 进GSK-3β表达，并抑制Akt/GSK-3β/CCND1/c-Myc信号通路来实现的。

Objective To explore the potential mechanism of Erzhi Wan in the treatment of multiple myeloma （MM）based on network pharmacology，molecular docking and cell experimental verification. Methods Network pharmacology and molecular docking were used to screen the key indicators of drugs-diseases-targets-pathways. MM line RPMI-8226 cells were cultured and given Erzhi Wan intervention. Apoptosis was detected by Annexin Vfluorescein isothiocyanate（FITC）staining. Transwell assay was used to determine cell invasion ability. Real-time quantitative polymerase chain reaction（qRT-PCR）was used to detect the mRNA expressions of protein kinase B （Akt），cyclin D1（CCND1），glycogen synthase kinase 3β（GSK-3β）and c-Myc. Western Blot was used to detect the protein expressions of Akt， phosphorylated protein kinase B（p-Akt）， CCND1， c-Myc， GSK-3β and phosphorylated glycogen synthase kinase 3β（p-GSK-3β）. Results Fourteen effective components of Erzhi Wan were obtained. There were 142 targets involved in the treatment of MM by Erzhi Wan. The results of pathway enrichment analysis showed that the key targets may be mainly concentrated in cancer pathways， lipids and atherosclerosis. Molecular docking results showed that luteolin and quercetin had good binding activity and stability with GSK-3β. Further cell experimental verification showed that compared with the blank group，the apoptosis rate of cells in the low-，medium- and high- dose groups of Erzhi Wan was significantly increased，the number of cell invasion was decreased，the mRNA and protein levels of GSK-3β were significantly increased，and the mRNA and protein levels of CCND1，Akt and c-Myc were significantly decreased in a dose-dependent manner，among them，the differences in the medium- and high- dose groups being statistically significant（P＜0.05 or P＜0.01）. Conclusion The therapeutic effect of Erzhi Wan on MM may be achieved by promoting the expression of GSK-3β and inhibiting the Akt/GSK-3β/CCND1/c-Myc signaling pathway through key active components such as luteolin and quercetin.

R285.5

广东省医学科研基金项目（编号：C2023074）；广东省中医药管理局项目（编号：20241093）