Study On Norcantharidin Chitosan Microspheres For Transcatheter Arterial Chemoembolization

Objective:To studied the norcantharidin-chitosan microspheres (NCTD-CS-MS) for transcatheter arterial chemoembolization (TACE) by using chitosan (CS) as carriers and norcantharidin (NCTD) as model medicine, based on the excellent plasticity of CS. Chemotherapeutic agents concomitantly administered remain in a tumor for a longer period at a higher concentration and selective arterial obstruction induces ischemic tumor necrosis while minimizing damage to the liver tissue using TACE for liver cancer in this study. To provide reference for clinical treatment of unresectalbe liver cancer.Method: (1) The NCTD-CS-MS were achieved by emulsification-chemical crosslink method and the technique conditions was optimized using central composite design(CCD). On the basis of single factor analysis, the yield, drug loading, envelop efficiency, mean diameter and span dispersity were used as observing indexs, and weight percent of NCTD and chitosan(X1), volume percent of organic phase and water phase(X2) and chitosan concentration in acetic acid water(X3) were used as factors while applying central composite design and response surface methodology (CCD-RSM) to optimize preparation process. (2)The particle size and distribution of microspheres were analyzed by optical microscope, and the surface characterization of the microspheres was identified with scanning electronic microscope (SEM). Application of infrared spectroscopy (FT-IR), X-ray powder diffraction (X-RD) and differential scanning calorimetry (DSC) to verify the formulation of chitosan matrix. Then the micromeritic characteristies, suspensibilit, fluidity and catheter bolus were considerated. (3) The release of MS prepared under different conditions in vitro was investigated with Dynamic dialysis method in pH 7.4 phosphate buffer solution (PBS) and the stability of microspheres was studied. (4) New Zealand rabbits were selected as model, embeded with VX-2 tumor tissue in the lobe of the liver using laparotomy or percutaneous liver paracentesis under ultrasonic guidance. Ultrasound and CT examinations were performed at different time periods after implantation respectively. (5) Animals which were inoculated of VX-2 liver tumor were divided into 4 groups randomly, and begun to be therapied at 3～4 weeks after inoculation. NCTD (1.0 mg·kg-1) was selected because of its known potency against VX-2 tumor. Group 1 was treated with TACE with NCTD-CS-MS(120～200μm, 10.09 mg·kg-1), group 2 was treated with intraarterial NCTD, group 3 was treated with intraarterial saline solution and group 4 was untreated. To observe the live time, the tumor volume and analyze tumor necrosis degree of every group, at the same time, the changes of hepatic function during pre-and post-treatment.Results: (1) The yield, drug loading, envelop efficiency, mean diameter and span dispersity were taken as indexes optimized by CCD-RSM , and the optimal condition was: the weight percent of NCTD and chitosan was 68.5 %( X1), volume percent of organic phase and water phase(X2) was 8 and chitosan concentration in acetic acid water(X3) was 3.2 %, while the envelop efficiency 32±0.83% and the drug loading 10.09±0.56 %. (2) SEM graph shows that microparticles were sphere, smooth, uniform and less of adhere. The average diameter was (143.54±4.24)μm and 87 % of the all microspheres in 60～200μm. FT-IR,X-RD and DSC graph indicated the reaction btween chitosan and glutaraldehyde, and NCTD dispersed in chitosan matrix. the micromeritic characteristies, suspensibilit, fluidity and catheter bolus were suitable. (3)The release of NCTD crossed by chitosan and glutaraldehyde followed Weibull distribution in vitro and was sustained for 15 d. The chitosan matrix maintained 2 months. (4) The study showed tumor grew productivly during 2～4 weeks after inoculation and tumor's body increased rapidly. 4 weeks later degeneration and necrosis at the center of tumor can be seen, peripheral lymph node lung metastasis were aslo be seen. Survival time of rabbits bearing carcinoma was 33.4±2.3days. Laparotomy had larger damage and recovering slowly to rabbit than percutaneous liver paracentesis. To synthetic compare, percutaneous liver paracentesis under ultrasonic guidance was employed. (5) After treatment, every 3, 7 and 14days, to evaluate the volume and growth rates of tumor with ultrasound: by measuring the diameter. The growth of tumor in microspheres group was inhibited continuously during the observation period of 14 d while there was a obvious inhibitin effect in NCTD group only at the first 3 d disappeared gradually thereafter. The microspheres showed a stronger inhibitin effect than that of NCTD and saline with statistic significance, while the difference between NCTD, saline and control has no statistic significance during the observation period of 7-14 d. Pathological examination showed that there was a great deal of coagulation necrosis in the tumors in the microspheres goups.Conclusion: The prepared NCTD-CS-MS have a high drug loading and entrapment efficiency, surface smooth and rounded. The extended release and chitosan matrix maintained 2 months can be reduce the toxicity of drugs and reduce the administration frequency. During TACE, the suspension and dispersion of microspheres were fit. The NCTD-CS-MS for hepatic artery embolization on VX-2 rabbit liver has a therapeutic effect.