Study On Docetaxel Submicroemulsion For Intravenous Injection

Docetaxel (Taxotere^TM is a new generation of anticancer drug. Similar to paclitaxel (Taxol^TM, it belongs to taxane and has been approved by FDA as taxane anticancer drugs. Docetaxel acts by disrupting the microtubular network that is essential for mitotic and interphase cellular functions. It promotes the assembly of tubulin into stable microtubules and inhibits their disassembly, causing inhibition of cell division and eventual cell death. The novel action mechanism and the unique efficacy in clinical trials lead to much wider application of docetaxel and its combination therapy in the treatment of many cancers, such as breast, non-small-cell lung, pancreas, soft tissue sarcoma, head and neck, stomach, ovarian, and prostate cancer etc.Because butoxy carbonyl group of docetaxel replaces the benzoic acid group of paclitaxel, the former's water solubility is relatively higher than that of the latter. Although possessing higher water solubility, the commercial product of docetaxel still adopted Tween 80 and ethanol as injection solvents. Taxotere^TM a brand name of docetaxel, used Tween-80 as solubilizer and ethanol as auxiliary solvent, which was packed separated as condensed injection solution and solvent in two packages. The condensed injection solution was formed by Tween-80 solution of docetaxel and the solvent was 13 % (w/w) ethanol. Before application, the solvent was firstly added into the condensed injection solution and mixed uniformly, then dilute it with 0.9 % NaCl solution or 5 % glucose injected solution before injection. The procedure of this formulation is complicated and very inconvenient for clinical use. Moreover, this procedure of dilution is prone to cause secondary contamination and therefore brings risks to the patients. Although, the formulation utilizes relatively less toxic solubilizer Tween 80 and ethanol as solvents, allergies are still seen in clinical application. Yet the incidence rate is lower than that of paclitaxel injection, the patients still have to receive anti-supersensitivity pretreatment before using it, in order to prevent supersensitivity side effect. Moreover, Tween 80 is prone to cause haemolysis. In order to diminish these problems of docetaxel formulation and overcome the low solubility of docetaxel, a docetaxel submicroemulsion for intravenous injection was developed in this study, so as to provide fundmental work for new dosage form research of docetaxel. The main contents of the research are as follows:1. Preformulation research of docetaxel submicroemulsion. A sensitive, precise and convenient HPLC method with high specificity for docetaxel determination has been developed, which was not only suitable for docetaxel content determination, but also suitable for the isolation of docetaxel and its degradation products or other impurities. Moreover, various external factors on docetaxel stability were investigated and the results indicated that the alkaline environment was the main influence on docetaxel stability, other influencing factors including high temperature, oxidation and light, while the influence of acid environment was relatively smaller.2. Formulation design of docetaxel submicroemulsion and the research of preparation techniques. Single factor analysis was employed to evaluate the influence of the concentration of phosphatidyl choline, Poloxamer 188 and the ratio between oleic acid and oil phase on the relevant properties of the submicroemulsion, respectively. Then based on the results of the single factor analysis, the formulation was optimized by orthogonal design and the stability parameter was taken as the response. In the preparation techniques research, the influences of emulsification methods, emulsification temperatures, emulsification time, homogenization pressure and homogenization times on the emulsion quality were investigated respectively and finally the optimal technical parameters were selected. The submicroemulsion prepared with the optimized formulation and preparation technique was stable with uniform particle size distribution and the dimension of 50% emulsion droplets was less than 169 nm.3. The stability study of docetaxel submicroemulsion. Extreme conditions, such as high temperature, freeze thawing, light and low temperature, were used to investigate the stability of docetaxel submicroemulsion. The results showed that high temperature and light would lead the drug content and pH to decrease. The particle size of the emulsion droplets increased significantly under freezing condition and demix of the emulsion was observed. However, the responses of docetaxel submicroemulsion didn't change significantly under low temperature. In addition, the results of physical stability of docetaxel submicroemulsion in different dilutions showed that the particle size in them didn't increase significantly within 5 h, which indicated that the preparation wouldn't likely cause clot in blood vessel after dilution.4. The pharmacodynamic study of docetaxel submicroemulsion. Tumor-loaded mouse model was established by subcutaneous injection of cancer cell. The anticancer effect of docetaxel submicroemulsion was investigated by measuring the changes of tumor volume, average tumor weights and tumor inhibition ratio, as well as observing the histopathological appearance of tumor tissues. The results indicated that the tumor tissues grew slowly after the administration of docetaxel submicroemulsion, and the growth of tumor volumes and weights was inhibited significantly. Histological results also showed that the growth of tumor tissues was inhibited and the necrosis was observed.5. The pharmacokinetics study of docetaxel submicroemulsions. A HPLC method was established to determine docetaxel in vivo. The plasma samples were dealt with liquid-liquid extraction method before determination and norethisterone was selected as the internal standard. This method was simple, quick, sensitive and very suitable for the drug determination in vivo. The pharmacokinetics results of docetaxel submicroemulsions in rats showed that AUC increased significantly (p<0.05) and the clearance rate (CL) was decreased (p<0.05), which illustrated that the docetaxel submicroemulsion could maintain a relatively higher concentration in blood and therefore enhancing anticancer effect.6. Preliminary safety evaluation of docetaxel submicroemulsion. The safety of docetaxel submicroemulsion was evaluated by abnormally toxic test, blood vessel irritation test and hemolysis test (common test tube method in vitro). The results showed that docetaxel submicroemulsions had neither serious intravenous toxicity nor undesirable irritation to veins. Moreover no hemolysis was observed in the common test tube method in vitro.7. The freezing drying study of docetaxel submicroemulsion. Lyophilization method was adopted to turn the docetaxel submicroemulsion into lyophilized emulsion. The formulation and the preparative technique of the lyophilized emulsion were investigated by studying the influences of the types and concentrations of the cryoprotectants, as well as the prefreezing temperature and rates on the relevant characteristics of the lyophilized emulsion. The results showed that trehalose, sucrose and maltose had better effects in five selected cryoprotectants, and the effect of maltose was the best. During prefreezing process, the temperature and rates had vital influence on the quality of the lyophilized docetaxel emulsion. Fast prefreezing with -45°C generated nice effects for minor change happened to the particle size of the reconstituted lyophilized emulsion.We have applied the relevant formulation and procedure of the docetaxel lyophilized emulsion established in this study for Chinese patents, which is under verification by relevant agency.