Polymeric Micelles as Carriers for a Hydrophobic Anti-cancer Drug

Abstract

Background and Aim: Polymeric micelles (PM) are used as drug carriers for three critical functions: drug solubilisation, drug targeting, and controlled release of drug. PM are attractive drug vehicles for anti-cancer drugs as they are nano-sized and easily prepared with good stability. SN25860 is a dinitrobenzamide mustard hypoxia-activated prodrug (HAP) with activity against solid tumours. However, this compound has poor aqueous solubility (intrinsic solubility of sodium salt is 50 μg/ml). The aim of this study was to develop a PM formulation based on Pluronic® copolymers to enhance the solubility of SN25860. The physicochemical properties of the delivery system were characterized for suitability as a parenteral formulation potentially for tumour-targeting. The safety was assessed through an in vitro haemolysis test. Methods: A rapid and reliable reversed phase HPLC method with UV detection at 360 nm was developed and validated for quantification of SN25860. Various PM platforms were prepared from two binary Pluronic® block copolymers systems, P123/F127 and P103/F127 using the solvent evaporation method. The platforms were characterized in terms of size, zeta potential, morphology, and physical stability over a 6 months period. SN25860 was loaded into the selected PM platforms and the changes in size, zeta potential, and morphology were investigated. The final formulation with high drug loading (DL) and entrapment efficiency (EE) was selected, and subjected to further studies; thermal analysis, in vitro drug release behaviour, and haemolysis test. Results and discussion: The HPLC method developed for determination of concentration of SN25860 comprised of an acetonitrile and Na2HPO4 buffer (50 mM, pH 3) (45:55, v/v) as mobile phase with a retention time of 6.2 min. The pH control was essential to maintain the SN25860, a weak acid with a pKa of 4.2, in unionised form for a satisfactory retention time. Formulations based on the P123/F127 systems have similar particle size but narrower size distribution compared with formulations based on the P103/F127 PM systems. The PM platforms were relatively stable in the first four months but the micelles started to degrade and became smaller at later time points. Formulations F2 and F4 based on P123/F127 were selected for encapsulation of SN25860 as they have moderate size, narrower size distribution and good stability. The average particle size formulation was smaller after the drug was loaded (~30 nm versus ~46 nm, p<0.01). Interestingly, with the same volume of hydration solution, increased the polymer concentrations (>48 mg/ml), the size of the blank PM was much smaller than that at lower concentration (~25 nm). For this formulation the size did not change significantly after drug loading (28 ± 2.7 nm) (p>0.05). For both F2 and F4, the EE was satisfactory (>90%) but the DL was lower (2%) compared to the literature. However a maximal drug concentration of 3 mg/ml was achieved which was sufficient for preclinical trial. The thermal analysis suggested that most of the drug was encapsulated into the PM. The comparison of freeze-dried formulation F2, and non-freeze-dried formulation F2 and F4 in drug release study indicated burst release in first few hours and delayed constant release for up to 24 hr. The haemolytic effect of the formulations was less than 10%, suggesting the formulations were biocompatible for parenteral delivery. Conclusion: The PM formulations are able to enhance the aqueous solubility of SN25860, from 50 μg/ml to 3 mg/ml. The drug-loaded formulation has a small size (30 nm), which is suitable for drug targeting (20 - 200 nm) and deep penetration into solid tumours. Even though the formulation has an initial burst release, the formulation is still able to maintain a sustained release for 24 hr. The drug demonstrated little or no haemolysis while the formulation F2 showed minimal haemolytic effect, thus F2 is a promising formulation for parenteral administration in preclinical trials.