Study On Preparation And Properties Of Paclitaxel-loaded Polymeric Micelles Prepared With Poly(2-ethyl-2-oxazoline)(PEtOz)and Poly(ε-caprolactone)(PCL)

Taxol(Paclitaxel), which is extracted from Taxus, represents a new class of anticancer agents and functions by promoting the assembly and stability of microtubules thereby causing mitotic arrest. Since be approved by FDA in USA to treat ovarian cancer in December1992, Taxol rapidly become incorporated into clinical cancer treatment and now is a mainstay in the treatment of cancers.Taxol has poor solubility in water owing to its complicated structure, and Cremophor EL is a necessary formulation vehicle when Taxol are used in clinic. However,Cremophor EL is not an inert vehicle, which exerts a range of biological effects, some of which have important clinical implications. Its use has been associated with severe anaphylactoid hypersensitivity reactions hyperlipidaemia, abnormal lipoprotein parerns, aggregation of erythrocytes and peripheral neuropathy. So new drug carriers for Taxol should be required for the safetyc linical use of Taxol.In this paper,a type of amphiphilic block copolymer composed of Poly(2-ethyl-2-oxazoline)(PEtOz) and poly(ε-caprolactone)(PCL) was synthesized. Poly(2-ethyl-2-oxazoline), as a hydrophilic part, is a non-toxic, non-immunogenic water soluble polymer. PCL, as a hydrophobic part, has biodegradability and can be degraded into glutamic acid which is a non-toxic, non-accumulative endogentic substance. Due to the amphiphilicity of this block copolymer, biorecognizable core-shell type micelles can be prepared in water. PCL which acts as a drug incorporation site form the inner-core of the structure. PetOz which form the hydrated outer shell, may cloak the hydrophobic core to avoid quickly entrapping by the RES and the increase the stability of the inner core. Due to the amphiphilicity of this block copolymer, biorecognizable core-shell type micelles prepared by a diafiltration method can be expected as suitable site-specific carriers of Taxol.Based on these aims, the drug-loaded polymeric micelles were evaluated in its properties, size and cell cytotoxicity, anti-tumor activity in vivo, pharmacokinetics of Taxol following intravenous administrations in rats. The results ar e as follows.1. Synthesis and Characterization of Poly(2-ethyl-2-oxazoline)-poly(s-caprolactone)(abbreviated as PetOz-PCL). PetOz-PCL was prepared by anionic polymerization of PetOz nitiated by PCL with different molar ratio of monomer/initiator. The products were characterized by IR、1H-NMR. The results showed that structure of PetOz-PCL was definite and the PCL segments existed in a-helical conformation.2. Self-assembling of Taxol/PetOz-PCL micelles ore-shell type micelles of the PetOz-PCL block copolymers were prepared by the dialysis method. UV and HPLC analysis indicated that hydrophobic drug was entrapped in copolymer micelles."With the increase in hydrophobic block content in ialysate, the solubilization of micelles were increasing. With an increase of the drug-fed amount, the drug loading coment was increased, but the entrapment efficiency was decreased.3. Measurement of cell membrane cytotoxicity of copolymers ccording to standard of ISO1993and GB/T16886documents, the cytotoxicity was evalutate with hemolytic activity assay. The result showed PEtOz-PCL block copolymers are safer than Tween80, a low molecular weight surfactant and a chitosan derivative, a polymeric surfactant in terms of the induction of membrane damage.4. The anti-tumor activity of Taxol/PetOz-PCL micelles in vitro. KB cells was cultured in vitro, and the cytotoxicity test was conducted with MTT assay. The result showed that when Taxol is in low concentration(<30μM/ml), the Taxol/PetOz-PCL micelles has less cytotoxicity than Taxol injections which use Cremophor EL as a vehicle (P<0.01).5. The anti-tumor activity of Taxol/PetOz-PCL micelles in vivo. A nude mice transplanted tumor model of human epidermoid carcinoma KB was founded. The anti-tumor activity of Taxol/PetOz-PCL micelles and Taxol injections with Cremophor EL were studied. The result showed that the tumor inhibiting rate of Taxol/PetOz-PCL micelles was superior to that of Taxol injections. Polymeric micelles based on amphiphilic block copolymers (PEtOz-PCL) of poly(2-ethyl-2-ox-azoline)(PEtOz) and poly(’-caprolactone)(PCL) were prepared. Paclitaxel was successfully loaded into PEtOz-PCL micelles using a dialysis method. The hydrodynamic diameters of paclitaxel-loaded micelles were in the range19.4-23.4nm with narrow size distribution. The higher the content of hydrophobic block in the block copolymers, the higher the loading efficiency for paclitaxel. The hemolysis test indicated that PEtOz-PCL micelles were safer as an injectable carrier than Tween80in terms of membrane toxicity, and showed membrane toxicity comparable to Cremophore EL. As judged by in vitro MTT assay, paclitaxel incorporated into micelles retained the inhibition activity for the proliferation of KB cells, and the inhibition activity patibility of docetaxel and paclitaxel in intravenous solutions was comparable to that observed with Cremophore EL-based commercial formulation. The estimation of loading capacity for paclitaxel and membrane toxicity, and proliferation inhibition activity for carcinoma cells suggest that PEtOz-PCL micelles may act as a useful carrier for paclitaxel. The biodistribution study of PEtOz-PCL micelles to estimate its function as a long-circulating carrier will be discussed elsewhere.