Research On Novel Formulations Of Docetaxel Including Cyclodextrin Inclusion Compound & Nano-eumulsions

Docetaxel is a milestone medicine in cancer therapy field. It is mainly used for the treatment of non-small cell lung cancer and breast cancer. However, its low solubility and bioavailability limit its clinical usage. As marketed docetaxel injection uses Tween-80 and ethyl alcohol as solubilizer, which has a slight hemolysis, and patients should take anti-histamine and glucocorticoids in advance, all these make its usage very inconvenient. According to the feature of docetaxel and clinic idea of cancer therapy, two suitable formulations and its preparation technology were screened out(including SBE-β-cyclodextrin inclusion compound and long circulation nanoemulsion). Refer to the marketed Docetaxel injection, the two formulation samples and a nanoemulsion formulation sample(already screened and prepared in lab) were compared, including the safety evaluation and stability in vitro and their pharmacokinetics characteristics.A new method for determination content of Docetaxel(HPLC isocratic elution) was developed and methodology validation was completed, its linearity、precision、specialization、limit of detection and limit of quantification were detected. The stability of docetaxel in hyperthermia, illumination, oxidation, alkali, acid and different pH conditions was investigated as well. The results of forced degradation test show that docetaxel is stable under illumination and oxidationcondition, relatively stable under the conditions of high temperature、strong acid,unstable under strong alkali condition. The results of stable test of docetaxel under different pH show that it is suitable for docetaxel under pH≤6.0 in preparation and storage process.Based on the solubility、compatibility and stability of pharmaceutic adjuvants, the best proportion of SBE-β-cycodextrin inclusion compound formulation was determined by UV mothed. The formulation was screened out by orthogonal experiment, and the result is 1:70. The suitable preparation process is dissolving docetaxel in 30 ml alcohol and then mixing with SBE-β-CD for 1h with the condition of 500 rpm, 25 ℃. The variation tendency of temperature and conductivity of formulations during freezing process were chosen as indicators, and the lyophilization process was selected by detecting its lowest eutectic point. The suitable lyophilization SBE-β-cycodextrin inclusion compound samples was screened out and verified by microscope 、 X-ray and DSC. The results of microscope show that the sample is round, and inclusion compound was formulated with the certification of X-ray and DSC.The solubility of API in different pharmaceutic adjuvants and the basic characteristics of these adjuvants, such as viscosity, were detected. On the base of single-factor experiment, the suitable adjuvants were seleted. Pseudo-ternary phase diagram was drawn as electric conductivity was chosen as assessment criteria. The suitable proportion of oil、water and emulsifier & co-emulsifier was confirmed. The proper ratio of drug-ester was determined as particle size and encapsulation efficiency were treated as assessment criterias. According to the Gouy-Chapman theory, the aqueous layer thickness was detected and then the content of DSPEPEG2000 was determined by measuring samples’ zeta-potential values. As to the preparation process of this formulation, high pressure jet homogenizer was used. The suitable homogenizer condition was selected by comparing the particle sizes. The suitable formulation of long circulation nanoemulsion is: Docetaxel 100 mg 、PEG400 4g 、 Lecithin 、 Solutol HS15 、 MCT 2g 、 DSPE-PEG2000 50 mg 、Mannitol 900 mg. The suitable preparation process is: mixing oil phase and water phase, high-speed shearing for 5min, high-pressure homogenizing 5 times under 530 Pa and then freeze-dried with lyophilizer. Observe the final samples and select appearance 、 dissolvability 、 particle size and entrapment efficiency as main evaluation indicators. The results show that sample’s apprearance is good and can be completely dissolved in 3s. The particle size is 53.3nm, with P.I. around 0.115. The entrapment efficiency is about 90.5%.Refer to the marketed Docetaxel injection; the release characteristics and cytotoxicity in vitro of these formulations were detected. Compared with marketed Docetaxel injection, these formulations all have some sustained releasing characteristics. Huvec cells were chosen to detect formulations’ vascular irritation in MTT experiment. The safety of these formulations from better to worse is inclusion compound, nanoemulsion, long circulation nanoemulsion and marketed Docetaxel injection with lower IC50 value.SD rats were chosen in this experiment. The suitable analytical method was developed and validated. Drugs were given through tail vein by injection and blood samples were drawn from orbit. The content of Docetaxel in blood was detected and DAS 2.0 pharmacokinetic procedure was applied to calculate relevant pharmacokinetic parameters. The results show that compared with marketed Docetaxel injection, inclusion compound formulation has significantly improved AUC, long circulation nanoemulsion and nanoemulsion formulations exhibit better sustained releasing characteistics, and their releasing time can prolong to 24 h.