Antitumor Activity And Structure-activity Relationship Of Seven Lanostane-type Triterpenes From Fomitopsis Officinalis And Fomitopsis Pinicola

Lanostane-type triterpenes(LTT)are antitumor activities that are found in the porous fungus generally. In this paper, structure, antitumor structure-activity relationship and anti-tumor mechanism of LTT were studied, including the following four parts:First, in order to obtain a more comprehensive understanding of effective LTT constituents from porous fungus, the structure and activities studies of LTT constituents from polyporaceace fungus had been carried out. So far, 159 kinds of LTT had been discovered from polyporaceac fungus, They have anti-tumor, anti-inflammatory, antibacterial and other pharmacological activities. However, the antitumor activity of LTT components separated from fungus had not been well studied. Such as, similar structure compound showed different effect to humor tumor line which position changed of LTT structure caused the differences of antitumor activity. How did the LTT compounds kill the tumor cells. Since the experiment in antitumor study had been carried out in vitro, further work was required to antitumor activity of the LTT compounds in vivo system.The fruiting bodies of F.officinalis and F.pinicola were used as materials to separate LTT constituents. The extracting and isolating process of the triterpene compounds from F.officinalis and F. pinicola were studied. The powder of F. officinalis fruit body and F. pinicola fruit body were extracted by petroleum ether(60-90℃)and ethyl acetate solvent gradient extraction method instead of others. Petroleum ether(60-90℃)and ethyl acetate extracts were isolated by silica-gel column chromatography with different ratio of solvent system, then received the gradient fluent with 20 ml ampoules rather than rough process. The LTT from The fruiting bodies of F.officinalis and F.pinicola got the highest yield. Seven LTT compounds were isolated and identified, they were(compound 1)Fomitopsin C(0.004%),(compound 2)3-Keto-dehydrosulfurenic acid(0.0964%),(compound 3)Dehydroeburiconic acid(0.1721%),(compound 4) 3-acetoxylanosta-8, 24-dien-21-oic acid(0.1043%),(compound 5)pinicolic acid A(0.0754%),(compound 6)trametenolic acid B(0.0065%),(compound 7)eburicoic acid(0.0087%).In vitro, Seven LTT compounds were evaluated against MCF-7, He La, Hep G2 and A549 cells lines by the standard MTT assay and the structure-activity relationship of antitumor activity was discussed. The test results showed that structure-activity relationship of LTT against tumor activity was obvious after replaced with acetoxyl or carbonyl at C-3 and hydroxy at C-15.H22 tumor mouse model was used to assay compounds 2, 3, 4 and 5 in vivo. The followingthree aspects were studied:(1) The influence of four kinds of monomer compounds in solid tumors itself were detected with measuring the anti-tumor rate, The morphological changes in H22 solid tumor cells by Hematoxylin-eosin(HE) staining, The expression of VEGF in tumor by ELISA. The results showed that compounds 2, 4 and 5 had significant antitumor activities at the dose of 20mg/Kg with and 65.31%, 56.71%, 58.72% suppression respectively approach to69.19% of CTX group in subcutaneous H22-implanted mice, The results of tumor biopsy showed that the tumor cells were deformed, necrosis and inhibited. Compounds 2 and 4 had obvious effect and the necrosis area and measurement were positively correlated. The results of detecting VEGF in tumor showed that compounds had inhibitor effect significantly to the expression of VEGF in tumor. Compound 2 and Compound 4 than compound 3 and compound 5.(2) The influence of four kinds of monomer compounds in immune system in H22 mice were detected with measuring the thymus index, spleen index and IL-2, IFN-γ, TNF-α and IL-4 levels. The results showed that The spleen index and the thymus of H22 mice were significantly higher than the normal group. Compared with model group, high and middle dose of compounds 2 and 3could decrease the spleen index, closing to the normal group, while low dose of compounds 2and 3 could increase the spleen index. Compared with model group, high middle and low dose of compounds 4 could decrease the spleen index, the low dose group(p<0.05). High and low dose of compounds 5 could increase the spleen index(p<0.05). The influence of Compounds 2,3, 4 and 5 on thymus index without regularity. The IFN-γ contents of increased significantly.The IL-4 contents of decreased significantly. The levels of IL-2 and TNF-α were not significantly correlated to tumor growth inhibition in mice.(3) Tested the influence of four kinds of monomer compounds in H22 mice were detected with measuring levels of MDA, SOD, CAT,and the contents of GSH-Px in blood serum. The results showed that The contents of GSH-Px increased significantly. The contents of MDA decreased significantly. The levels of SOD and CAT were not significantly correlated to tumor growth inhibition in mice.