An Examination Of The Effects Of Lipids、P-gp Inhibitor And Self Micro-emulsifying Drug Delivery Systems On The Intestinal Lymphatic Transport Of Paclitaxel

Paclitaxel (PTX) is an important antineoplastic agent that has a wide range of effects on the treatment of various cancers including ovarian cancer, breast cancer and non-small cell lung cancer, colon cancer, head and neck cancer, lymphoma and AIDS-related Kaposi sarcoma. Currently, PTX formulations available for clinical use (6 mg-mL"’) are formulated in a mixture of ethanol and polyoxyethylated castor oil (Cremophor EL) (1:1 by volume) and are administered through intravenous (i.v.) infusion. However, the vehicle used in formulation, Cremophor EL, may contribute to the hypersensitivity reaction and neurotoxicity in patients, which are notorious side effects associated with the use of i.v. PTX. Thus, much research is being carried out to develop oral formulations of PTX to minimize the i.v. route-related adverse effects. On the other hand, oral administration is hampered by the low bioavailability due to PTX’s low solubility in water; cytochrome P-450 enzymes in the gut and liver, and high affinity to the multidrug efflux pump P-glycoprotein (P-gp). Lipids formulation could offer an opportunity to enhance the oral bioavailability of paclitaxel. Mechanisms by which lipids improve the bioavailability of PTX may include enhanced solubilization of PTX in the gastrointestinal lumen and increased absorption via selective lymphatic uptake. However, the degree of intestinal lymphatic transport of paclitaxel in rats has not been published previously and remains to be validated by experiment. The current study has therefore investigated the lymphatic transport of paclitaxel in an anesthetized mesenteric-lymph-duct rat model. The study was divided into three parts, detailed as follows:Part Ⅰ:A sensitive high-performance liquid chromatographic (HPLC) method was developed for the determination of paclitaxel in micro-samples of rat plasma and rat lymph in order to study the lymphatic transport of paclitaxel. The HPLC system consisted of a Waters 2695 Alliance module and Waters 2487 dual λ absorption detector. A Waters Symmetry(?) C18 analytical column (150 × 4.6 mm,5 μm) and a C18 guard column (Security Guard, C18,4 mm × 3 mm id; Phenomenex) were used for separation. The mobile phase consisted of acetonitrile:water (48:52, v/v) at a flow rate of 1 mL/min. The temperature of the column was set at 30℃, and the UV detector was set at 227 nm. The injection volumes of the lymph and plasma sample were 20 μL and 50 μL, respectively. The detection limit of PTX in the lymph was 50 ng·mL-1; the range of linear response for the lymph sample was 0.05～10 μg·mL-1 (r=0.9996). Recoveries of PTX in lymph fluid at concentrations of 0.05,0.5, and 5 μg·mL-1 were (78.1±4.65)%, (86.1±9.56)%、(85.2±5.19)%, respectively. The intra- and inter-day variabilities of paclitaxel were both less than 15%. The detection limit of PTX in the plasma was 20 ng·mL-1; the range of linear response for the plasma sample was 0.02～5 μg·mL-1 (r=0.9999). Recoveries of PTX in plasma at concentrations of 0.05, 0.5, and 2μg/mL were (77.3± 5.61)%、(77.0±2.33)%、(83.1± 5.79)%, respectively. The intra- and inter-day variabilities of paclitaxel were both less than 10%. This validated method for the assay of paclitaxel in micro-sample rat plasma and lymph is simple, sensitive and accurate, and can be used for pharmacokinetics studies of PTX in rats.Part Ⅱ:An anesthetized, mesenteric lymph duct-cannulated rat model was developed for the study of intestinal lymphatic transport of PTX. PTX solution alone, PTX solution pretreated with the p-glycoprotein inhibitor verapamil, and/or PTX and a 2:1 (w/w) mixture of linoleic acid:glycerol monooleate were administered intraduodenally to anesthetized rats. The extent of lymphatic transport of PTX over 8 h was (0.038±0.003)% after intraduodenal administration of PTX solution. Coadministration of a mixture of linoleic acid-monoolein significantly increased the extent of intestinal lymphatic transport of PTX, but it had little impact on the absolute oral bioavailability of PTX. In contrast, pretreatment with verapamil increased both the extent of lymphatic transport (3.5-fold) and absolute oral bioavailability (1.8-fold). Further increase in the lymphatic transport (6.5-fold) and absolute oral bioavailability (1.8-fold) was achieved by the combination of pretreatment with verapamil and coadministration with the linoleic acid-monoolein mixture. These data indicate that the application of lipid vehicle holds promise for selectively targeted lymphatic delivery of PTX. P-glycoprotein inhibition can result in both increased intestinal lymphatic levels and absolute oral bioavailability of PTX.Part Ⅲ:A new, self micro-emulsifying drug delivery system (SMEDDS) of PTX was developed to study the lymphatic transport of PTX in the mesenteric lymph duct-cannulated anesthetized rat model, and the extent of lymph transport of PTX was compared with PTX solution. Pseudo ternary phase diagrams were applied to identify the efficient self-emulsification region and optimize the formulation of SMEDDS. A pharmacokinetic study was conducted in the mesenteric lymph duct-cannulated anesthetized rat model to assess the lymph exposure after an intraduodenal paclitaxel dose of 20 mg·kg-1 in the SMEDDS formulations. The extent of lymphatic transport of PTX over 8h was (0.151±0.022)% after intraduodenal administration of PTX-SMEDDS formulation, which is significantly increased compared with the control group. The Cmax and AUC of SMEDDS were higher than those of control group, and the bioavailability was significantly increased. These results suggested that the prepared SMEDDS formulations produced acceptable properties in terms of lymphatic transport and could increase the bioavailability of paclitaxel.The objective of this study was to further clarify the mechanism of oral gastrointestinal absorption of PTX and provide some information for the research of PTX new formulations.