| Water decoctions from the leaves of Taxus are used in traditional Chinese medicine to treat cancer, indicating that water soluble constituents of this tree may indeed possess anticancer activity. In an effort to search for natural products from Taxus with improved pharmacological features compared to paclitaxel,7-xylosyl-10-deacetylpaclitaxel has been isolated and identified by our group. This is a naturally occurring xyloside, which has been shown to possess higher water solubility than paclitaxel. However, preliminary studies suggested that 7-xylosyl-10-deacetylpaclitaxel was not significantly absorbed after oral administration, and pharmacokinetic studies yielded low plasma concentrations of the active ingredient. It is necessary to investigate whether the observed low oral bioavailability is due to poor absorption. Therefore, we studied the transepithelial flux of 7-xylosyl-10-deacetylpaclitaxel using the human colonic cell line Caco-2, a well established model of human intestinal absorption, and the results showed that:1. Caco-2 cell model was authenticated by determination of TEER of Caco-2 cell monolayers.The results indicated that in our laboratory, Caco-2 cells had reached integrated confluence and formed differentiated mono layers 21 days after seeding. TEER of the cells reached 300Ωcm2 and met the requirement of drug transport. So this model can be an in vitro model that investigated intestinal permeability and transport of 7-xylosyl-10-deacetylpaclitaxel.2. Methyl thiazoly tetrazolium chromatometry was applied to study the cytotoxicity of 7-xylosyl-10-deacetylpaclitaxel in Caco-2 cells.The results showed that compared with the control group,drug solution concentration lower than 20μM had no toxicity to Caco-2 cell.while drug solution concentration higher than 20μM was toxical to Caco-2 cell.3. The flux of 7-xylosyl-10-deacetylpaclitaxel (0.5-20μM) across the Caco-2 cell layer was linear with time for up to 3 hr. The data indicated that 7-xylosyl-10-deacetylpaclitaxel was primarily absorbed by passive diffusion across the Caco-2 cell membranes.4. The Papp for BL to AP transport decreased from 25.8±6.3×10-6 cm/sec (mean±SE) at 0.5-2μM to 17.1±3.8×10-6 cm/sec at 5-20μM 7-xylosyl-10-deacetylpaclitaxel. The Papp for AP to BL transport was fairly constant (16.3±6.3×10-6) throughout the concentration range studied. Hence, the observed BL to AP flux was the result of passive diffusion plus active transport. The AP to BL flux may be the result of passive minus active transport.5. Verapamil and tetrandrine reduced the efflux by about 50%. Concomitant with this inhibition we observed a about 2.5-fold increase in the AP to BL flux. It was shown that 7-xylosyl-10-deacetylpaclitaxel is a substrate of P-gp. 6. The apparent KM value of 93.4μM for the saturable process for 7-xylosyl-10-deacetylpaclitaxel was more than that previously reported for the P-gp substrate paclitaxel (16.5μM) in the Caco-2 cell system. The Vmax value of 909.1 pmol/hr/cm2 for 7-xylosyl-10-deacetylpaclitaxel was similar to that calculated for paclitaxel (1050 pmol/hr/cm2).This indicated that the active transport rate of 7-xylosyl-10-deacetylpaclitaxel was similar to that of paclitaxel.7. This was 16.3×10-6 cm/sec for 7-xylosyl-10-deacetylpaclitaxel and only 4.4×10-6 cm/sec for paclitaxel. According to previous studies, a Papp value in Caco-2 cells of>10×10-6 cm/s suggest good drug absorption in vivo corresponding to 70-100% of the applied dose.In summary, it is hypothesized that the low oral bioavailability of 7-xylosyl-10-deacetylpaclitaxel may be more dependent on presystemic metabolism in the liver than on lack of absorption.
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