Studies On The Preparation Technology Of Temsirolimus-loaded Liposomes

Temsirolimus is C-42hydroxyester of sirolimus, which belongs to the mTOR kinase inhibitor class of antiproliferative agents. Temsirolimus is able to bind the cytoplasmic protein FKBP12, then the temsirolimus-FKBP12complex inhibits the activity of mTOR kinase, followed by inhibition of multiple key signal transduction pathways and blocking cell cycle from the Gl to S phase, thereby playing role of anticancer. Widespread clinical studies indicates that Temsirolimus has a profound impact in the treatment of variety of cancers such as renal cell carcinoma, breast cancer, endometrial cancer, non-small cell lung cancer, etc. Because of its poor aqueous solubility, Temsirolimus is formulated as injections using Tween-80and PEG400as solubilizer. However, this commercial formulation is poor stable and often leads to hypersensitivity reactions, meanwhile the non-selective distribution of drug results in dose-limiting adverse reaction during clinical application.This paper aims to develope new drug delivery system which would improve the solubility of drugs, reduce adverse reaction and facilitate clinical applications. Liposome has numbers of advantages as an excellent formulation for anticancer drugs. In present work, we selected phospholipids and cholesterol to encapsulate Temsirolimus and prepared lyophilized Temsirolimus liposomes. The main methods and results were as follows:The dialysis method was applied to separate the free drug from drug-loaded liposome. HPLC method was established to determine the content of drug and the entrapment efficiency of liposome, which was proved to be of high accuracy and good reproducibility.Thin-film dispersion method was employed to prepare Temsirolimus liposomes, Influential factors such as temperature and ultrasonic time were investigated, and then optimum procedure of preparation was obtained. Single factor experiments were conducted to evaluate the effect of ratio of phospholipids to drug, the ratio of phospholipids to cholesterol and the concentration of phospholipids on EE.Response surface methodology was used to optimize the Prescription, i.e. the ratio of phospholipids to drug, the ratio of phospholipids to cholesterol and the concentration of phospholipids was20:1,14:1,3%respectively. Under these conditions, the EE was maintained to83.0%, which had only0.35%relative error compared with predictive value.Freeze drying technique was utilized to prepare lyophilized liposomes. Influential factors such as freezing temperature, freezing time and drying time were investigated, and then optimum procedure of lyophilized was obtained. Taking shape, reconstruction effects and changes in particle size as indices, sucrose and mannitol were selected as cryoprotectants and the ratio of sucrose to mannitol was1:1.Finally, the character of Temsirolimus liposomes before and after lyophilized was studied, then the quality and stability of liposomes were initially evaluated. Under the optimum conditions, Temsirolimus liposomes obtained were uniformly spherical in shape, and the particle size as well as encapsulation efficiency of liposomes changed little after lyophilized. The MTT assays were performed to evaluate the influence of Temsirolimus liposomes on HepG2cells viability.