PH-sensitive Poly(Ethylene Glycol)-poly(Lactic Acid)-poly(β-amino Ester) Copolymer Micelles For Paclitaxel Delivery

Objective:Taxol (paclitaxel, PTX, commodity name Taxol), is a kind of highly effective cytotoxic drugs by stabilizing microtubules, inhibiting cell mitosis and induction of cell apoptosis. Paclitaxel was approved by the FDA in December,1992 for the treatment of advanced ovarian cancer drugs, as well as for the treatment of breast cancer thereafter. At present, paclitaxel has been extensively used for treatment of breast cancer, non-small cell lung cancer, ovarian cancer, kaposi’s sarcoma, and a variety of malignant tumor, which has been regarded as one of the first class anti-cancer drugs in the world.The physic-chemical properties of taxol showed that its solubility in water is as low as 0.25 mg-L"1, hence it has been a restrictive factor for its extensive application owing to the low solubility in aqueous milieu. In the commercialized product taxol injection, paclitaxel was solubilized by polyoxyethylenated castor oil (Cremophor EL)/ethanol (50:50) which is mainly used for the treatment of ovarian and breast cancer. However a large number of studies have shown that paclitaxel injections solubilized by cremophor EL are prone to cause side effects such as allergic reaction, renal toxicity, neurotoxicity, respiratory depression, sleepiness, and low blood pressure, which has greatly restricted its clinical application. Hence, nanoparticles such as liposomes and micelles, etc, have been applied to increase the solubility of the drug, due to the oil injection of paclitaxel has not been approved for clinical setting.By consideration of the aforementioned factors, the thesis employed poly(ethylene glycol)-poly(lactic acid)-poly(β-amino ester) block polymer as the drug carrier material, to increase its solubility with reduced side effects. The copolymer can form polymer micelles in water at concentrations above of its critical micelle concentration (CMC), with a hydrophobic core composed of poly(lactic acid) and poly(β-amino ester),and a hydrophilic corona of poly(ethylene glycol) (PEG).The hydrophobic drug paclitaxel can be solubilized into the hydrophobic core. The poly(β-amino ester) moiety would spread under acidic conditions, with the induction of the fast release of the encapsulated drug.The synthesis of poly(ethylene glycol)-poly(lactic acid)-poly(β-amino ester) block copolymer and preparation of paclitaxel-loaded micelles with enhanced solubility in aqueous milieu can be referred below.Method:This part is composed by the preparation of pH sensitive amphiphilic block polymer, controlled drug entrapment and release, as well as in vitro and in vivo performance of drug loaded micelles.(1) PBAE was synthesized by Michael addition reaction. The structure and molecular weight of PBAE was characterized by 1H NMR. The pKb value was determined by acid-base titration. The critical micelle concentration (CMC) of the copolymers was measured by pyrene fluorescence spectroscopy.(2) Paclitaxel (PTX) loaded copolymer micelles were prepared by membrane hydration methods. Drug loading efficiency (DL) and entrapment efficiency (EE) of micelles were evaluated by high-performance liquid chromatography (HPLC). The size of micelles was determined by nanoparticle size analyzer. In vitro release was performed in phosphate buffered saline(pH 7.4,pH 6.5 and pH 5.5) at 37℃.(3) MTT method was used to evaluate the toxicity of PBAE and PELA to MCF-7 breast cancer cells under different pH conditions.(4) The biodistribution of taxol injection, PELA-PTX, PBAE-PTX was compared in rodents through i.v. administration.Result:(1) The results indicated that the pKb values of PELA and PBAE are 7.1 and 6.5 respectively. The CMC of the above polymers are 3.31 mg·L-1 and 4.79 mg·L-1, respectively.(2) No big difference was observed in the drug loading amount and encapsulation efficiency of the two materials. The drug loading amount of the micelles were 12.37% and 12.05%, respectively, and the entrapment efficiency were 70.55% and 68.53%, respectively. The paclitaxel loaded micelles were stable within a week at 4℃. In vitro release study showed that the drug release from PELA micelles increased slightly with the decrease of the pH in release milieu. However, the drug release accelerated obviously upon acidic environment (pH 6.5 and pH 5.5) from pH responsive polymer micelles. The released drug after 24 h increased from 65% to nearly 80%, at pH 7.4 and pH 5.5, respectively.(3) Cell toxicity experiments show that under the condition of different pH value, PELA and PBAE polymer nano micelles are non-toxic to breast MCF-7 cell under the concentration range, at the same time, in high concentration PBAE on MCF-7 cell toxicity are increased with the decrease of the pH, PELA have no obvious change.(4) Paclitaxel injection drug volume in various organizations from high to low order as:liver> spleen> lung> kidney> heart> blood; And PELA-PTX drug volume in various organizations from high to low order as:lung> liver> kidney> spleen> heart> blood, PBAE-PTX drug accumulation amount in each group has a high to low is:the lung> liver> kidney> spleen> heart> blood, PELA-PTX, PBAE-PTX show long cycle effect and obvious EPR effect.Conclusion:Based on the above results, we concluded that the as-prepared block copolymer micelles showed high performance in increasing paclitaxel solubility, high drug loading and long stability, as well as pH controlled drug release from micelles. Such pH sensitive polymer would find promising application in anti-cancer drug delivery can obviously increase the paclitaxel solubility in water, and the drug-loading efficiency is high and stable, the polymer are promising smart carriers for anti-tumor drugs with the advantages of pH-responsive drug release.