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研究発表を行った学会；
・The 6th Asian Cyclodextrin Conference
　２０１１年８月２８日〜３０日（キャンベラ、オーストラリア）

タイトル；Preparation and Evaluation of Folate-appended Methyl-β-Cyclodextrin as New Antitumor Agents.

発表者；小野寺　理沙子　氏
　　　（熊本大学　大学院薬学教育部　製剤設計学分野）
Abstract；
【Purpose】　Methyl-β-cyclodextrin (M-β-CyD) is widely used as a raft disrupting agent through extraction of cholesterol from cholesterol-rich lipid rafts which are abundant in tumor cell membranes1. Grosse et al. reported that intraperitoneal administration of M-β-CyD had antitumor activity in mice2. Therefore, M-β-CyD may have the potential as new antitumor agents which target cell membranes. On the other hand, the widespread use of the folic acid (FA) as a tumor-targeting ligand has been known, because folic acid receptor (FR) overexpresses in various kinds of epidermal tumor cells. Therefore, in the present study, to obtain more potent tumor cell-selectivity and antitumor activity of M-β-CyD, we had newly prepared folate-appended M-β-CyD (FA-M-β-CyD), and evaluated in vitro and in vivo antitumor activity of FA-M-β-CyD.
【Methods】 FA-M-β-CyDs were synthesized through tosylation and amidation into one of primary hydroxyl group of M-β-CyD before addition of FA by condensation reactions. The degrees of substitutions of FA of FA-M-β-CyD were determined by fast atom bombardment (FAB)-Mass and 1H-NMR. Cytotoxic activity of FA-M-β-CyD and FA-M-β-CyD/doxorubicin (DOX) complex in KB cells (FR positive; FR (+)) and A549 cells (FR negative; FR (-)) was determined by the WST-1 method. FR-mediated cellular uptake of FA-M-β-CyD was confirmed by a fluorescent spectroscopy. In vivo antitumor activity of FA-M-β-CyD was evaluated after intratumoral injection to BALB/c mice bearing Colon-26 cells.
【Results and discussion】 The 1H-NMR study demonstrated that FA-M-β-CyD having the average degree of substitution of FA of 1 was prepared. FA-M-β-CyD showed significant cytotoxic activity for KB cells (FR (+)), not A549 cells (FR (-)), in a dose-dependent manner. Cytotoxic activity of FA-M-β-CyD for KB cells was suppressed by the addition of FA as a competitor of FR. The fluorescence intensity of FA-M-β-CyD in KB cells (FR (+)) was significantly higher than that in A549 cells, suggesting that the cellular uptake of FA-M-β-CyD mediated FR-dependent endocytosis. FA-M-β-CyD enhanced cytotoxic activity of DOX in KB cells (FR (+)). FA-M-β-CyD showed the inhibitory effect on tumor growth in BALB/c mice bearing Colon-26 cells after intratumoral injection. These results suggest that FA-M-β-CyD can be the potential as not only a new antitumor drug but also a DDS carrier for DOX.

(1) Galbiati F.; Razani B.; Lisanti M P., Emerging themes in lipid rafts and caveolae. Cell, 2001, 106, 403-411.
(2) Grosse P.Y.; Bressolle F.; Pinguet F., Antiproliferative effect of methyl-β-cyclodextrin in vitro and in human tumour xenografted athymic nude mice. Br. J. Cancer, 1998, 78, 1165-1169.