Preparation And Evaluation Of Goserelin Microspheres-Gel Multiple Sustained-Release System

Goserelin is a decapeptide analogue of gonadotropin-releasing hormone.Clinically,it has a significant therapeutic effect on hormone dependent diseases such as prostate cancer and breast cancer.Poly（lactic-co-glycolic acid）（PLGA）microspheres can be used as the carrier of goserelin to achieve long-term sustained-release delivery of drugs.As a water-soluble drug,goserelin tends to migrate from the inner water phase to the outer water phase with the aqueous solution during the preparation process,forming a low encapsulation efficiency.In addition,due to the release mechanism of PLGA microspheres,the drugs will be released in a three-phase release pattern,namely,the initial burst release,the subsequent delayed release and the final rapid release of drugs.The fluctuation of blood drug concentration caused by the three-phase drug release mode may lead to the occurrence of adverse reactions.Therefore,we need to develop a microsphere system that can overcome the above obstacles.In this study,goserelin microspheres were prepared by the double emulsion solvent evaporation method with the internal water phase of poloxamer gel,and dispersed in the poloxamer gel matrix to construct a goserelin microspheres-gel multiple sustained-release system to modify the three-phase drug release mode and improve the entrapment efficiency.The optimal prescription process of goserelin microsphere was selected by single-factor analysis using the drug loading,encapsulation efficiency,particle size and in vitro release as the evaluation indexes;the prescription of thermosensitive gel was optimized by central composite design,and its viscosity and gelling temperature were evaluated.The optimized microspheres and gel were used to construct a goserelin microspheres-gel sustained-release system,and its in vitro release and in vivo pharmacokinetics were investigated.The specific results of this study are as follows:1.Preformulation study of goserelin microspheres preparationThe basic physicochemical properties of goserelin were studied,and a HPLC method for the determination of goserelin in microspheres was developed.The method was validated in terms of linearity,specificity,precision and accuracy,and the method was found to have good accuracy in the range of 0.01～1 mg·m L^-1.2.Preparation and evaluation of poloxamer thermosensitive gelThe formulation of thermosensitive gel was optimized by central composite design:the mass fractions of PL188 and PL407 were 2.39%and 23.37%,respectively.The prescription was investigated to have a gelation temperature of 36.9℃,and a viscosity of89.95 m Pa·s at 25℃,with good syringeability property and gel transformation at body temperature.3.Preparation and prescription process optimization of goserelin microspheresGoserelin microspheres were prepared by double emulsion solvent evaporation method,and the optimal prescription process was selected through single factor analysis:The internal water phase is a thermosensitive gel with 2.39%PL188 and 23.37%PL407,the concentration of PLGA 5050 2A is 250 mg·mL^-1,the concentration of PVA is 1%,the oil/water phase ratio is 1:80,the content of acetic acid in the internal water phase is 1%,the solidification temperature is 30℃,and the homogenization speed is 4000 rpm.The microspheres prepared from this prescription were validated to have a drug loading of3.70%,an encapsulation rate of 89.76%,and a particle size of 49.9μm.Circular dichroism and infrared spectroscopy showed that the structure of goserelin remained intact and unchanged in the microspheres.4.Preparation and in vitro evaluation of goserelin microspheres-gel sustained-release systemThe goserelin microspheres-gel multiple sustained-release system was established and its release in vitro was investigated.The results showed that the system had no burst release,short lag time,and its release period was about 18 days longer than that of placebo microspheres.The release period was shortened from 50 days to 20 days by using PBS（p H7.4）buffer containing 5%methanol at 45℃ as the accelerated release condition in vitro.The accelerated release curve correlated well with the in vitro long-term release curve after fitting analysis.5.In vivo study of goserelin microspheres-gel sustained-release systemThe plasma drug concentration of goserelin in rats was determined by ELISA method.Based on the plasma drug concentration curve and pharmacokinetic parameters,it was found that the goserelin microspheres-gel multiple sustained-release system had a relatively stable blood concentration in vivo with a C_max of 2.4 ng·mL^-1,and a release time of about50 days.HE staining analysis showed good biocompatibility of the system in rats.Summarize the above,the microspheres-gel multiple sustained-release system established in this study can achieve stable long-acting sustained-release delivery of goserelin with high drug loading capacity,which can provide an option for the long-acting effect of protein/peptide drugs.It is a promising long-acting sustained-release delivery system.