Abstract: Objective To explore the mechanism of Suanzaoren-Yuanzhi drug pair in the treatment of breast cancer related insomnia(BCRI)based on network pharmacology and animal experiments. Methods The core targets of Suanzaoren-Yuanzhi drug pair for treating BCRI were screened through TCMSP database, HERB database, and DisGeNET database. The STRING database was applied to construct an interaction network diagram of the core target proteins for the treatment of BCRI with Suanzaoren-Yuanzhi drug pair. The core targets were subjected to GO and KEGG enrichment analysis through the Metascape platform. Sixty mice were divided into control group, BCRI group, Suanzaoren-Yuanzhi drug pair group, diazepam group, Suanzaoren-Yuanzhi drug pair+LY294002(PI3K inhibitor)group, with 12 mice in each group. The control group mice were only used to establish a breast cancer model, without undergoing chronic unpredictable stress or intraperitoneal injection of cyclophosphamide. In other groups, the BCRI models were established using the method of breast cancer modeling + chronic unpredictable stress + intraperitoneal injection of cyclophosphamide. After successful modeling, administration was performed once daily for 7 consecutive days. The latency and duration of sleep in mice were detected. HE staining was used to detect hypothalamic pathology. ELISA was used to detect the levels of 4-aminobutyric acid(GABA)and 5-hydroxytryptamine(5-HT)in the hypotha-lamus. QRT-PCR was used to detect the mRNA expressions of brain and muscle arnt-like 1(BMAL1)and circadian locomotor output cycles kaput(CLOCK)in the hypothalamus. The phosphorylated phosphatidylinositol 3-kinase(p-PI3K)and phosphorylated protein kinase B(p-AKT)proteins in the hypothalamus were detected. Results The core targets for the treatment of BCRI using Suanzaoren-Yuanzhi drug pair were AKT1, TP53, TNF, ALB, HIF1A, STAT3, ESR1, BCL2, HSP90AA1, CASP3, PPARG, HSP90AB1, MAPK3, TGFB1, MMP9, MTOR, and CCND1. GO enrichment analysis for the aforementioned 17 core targets revealed that the cellular component(CC)of Suanzaoren-Yuanzhi drug pair for treating BCRI comprised 23 entries, the molecular function(MF)comprised 47 entries, and the biological process(BP)comprised 211 entries. KEGG enrichment analysis identified 116 pathways, predicting that the PI3K/AKT signaling pathway might be a crucial mechanism for Suanzaoren-Yuanzhi drug pair in the treatment of BCRI. Animal experiments indicated that, compared with the control group, the pathological damage to the hypothalamus of mice in the BCRI group was severe, the sleep latency extended, the duration of sleep decreased, the levels of GABA and 5-HT decreased, the expressions of BMAL1 and CLOCK mRNA, and p-PI3K and p-AKT proteins in the hypothalamus were reduced(all P<0.05). Compared with the BCRI group, the pathological damage to the hypothalamus of mice in the Suanzaoren-Yuanzhi drug pair group and diazepam group was reduced, the sleep latency shor-tened, the duration of sleep increased, the levels of GABA and 5-HT incresed, the expressions of BMAL1 and CLOCK mRNA, and p-PI3K and p-AKT proteins in the hypothalamus were elevated(all P<0.05). LY294002 reversed the effects of Suanzaoren-Yuanzhi drug pair on hypothalamus pathology, neurotransmitter and circadian rhythm in BCRI mice. Conclusion Suanzaoren-Yuanzhi drug pair can improve the hypothalamic pathology of BCRI mice, promote neurotransmitter balance, restore circadian rhythm, and improve sleep. Its mechanism of action may be related to the regulation of the PI3K/AKT pathway.

Key words: Suanzaoren-Yuanzhi drug pair, Breast cancer related insomnia, Neurotransmitters, Circadian rhythm