Bioactive triterpenes of jujube in the prevention of colorectal cancer and their molecular mechanism research

Jujube, a popular fruit from the Rhamnaceae family, relieves colorectal inflammation caused by spleen deficiency and has been used in many formulas in clinical for decades to treat colorectal cancer (CRC). As of yet, the therapeutic substances and mechanism of their action are unknown.The purpose of this study is to define the therapeutic substances of jujube and its mechanism of action in treating CRC.The pharmacological effects of jujube extract and its fractions were evaluated in vivo using a CRC mouse model induced by AOM/DSS. The DAI value, colon length, mortality, tumor burden, and histological tumor size of the treated animals were compared. To explore the potential therapeutic substances, LC-MS analysis was conducted to characterize the serum migration components. A network pharmacology experiment was carried out for potential molecular targets. To verify the therapeutic substances as well as the molecular mechanism of jujube intervening CRC, cellular MTT assay of the serum migration components, Western blot and IHC tests were conducted.The in vivo pharmacological studies showed that compared to AOM/DSS treated mice, the mortality and DAI value, tumor burden, and histological tumor size of jujube extract and its fat-soluble fraction (mainly contained triterpenes) treated mice were significantly reduced, and their colon lengths were obviously longer than AOM/DSS treated mice. The targeted-LC/MS analysis supposed triterpenes 3, 7, 9, 11, 12, 14, 17 - 21, and 25 - 28 to be the serum migration components, which might be the potential therapeutic substances. In the network pharmacology experiment, the GO annotation and enrichment analysis of the KEGG pathway indicated that PI3K-Akt pathway and inflammatory reaction were important factors for jujube inhibiting CRC. Cellular MTT assay of serum migration components indicated that the potential effective substances from fat-soluble fraction to be triterpenes 3, 7, 17, 19, 20, and 25. The Western blot and IHC assays implied that the jujube extract, its fat-soluble fraction, and triterpenes 7, 17, and 20 showed inhibition on the expression of PI3K/Akt/NF-κB signaling pathway-related proteins. Additionally, it was noted in the pharmacodynamic experiment that ZJL's effectiveness was more apparent than ZJH and SQL in tumor burden rate, colon length, and spleen weight, which indicated that the efficacy of ZJ is contributed from CD and SQ, and they may have a synergistic effect on anti-CRC.These results for the first time provide evidence that jujube triterpenes possess an anti-CRC effect, their mechanism was involving the control of the PI3K/Akt/NF-κB signaling pathway. What's more, the potential synergistic effect of the fat-soluble and water-soluble components found in this study provided a solid foundation for our deep understanding of how jujube can ameliorate CRC.