Category: Formulation and Quality
Purpose: Recently, developments of nanomedicines have been growing all over the world. Nanomedicines are able to improve efficacies and safeties, but they are more complex in terms of their functions and structures than traditional low molecular weight compounds. Due to their complexities, some guidelines for nanomedicines were published from the authorities, such as FDA, EMA and PMDA. In the guidance, detail characterization studies should be conducted for developing nanomedicine and developers should be justified their critical material attributes and critical quality attributes through the characterization studies.
In this study, polyethyleneglycol-polylactide block copolymer (PEG-PLA) which hydrophobic domains attributes different crystallinity to their isomer were chosen as a model polymeric micelle. Tamoxifen (TAM), a drug for the breast cancer was chosen as a cargo for polymeric micelles. The aim of this study is comprehensive characterization studies how the PLA crystallinity and PLA molecular weight of PEG-PLA block copolymer affects the loading or releasing TAM. In addition to the conventional characterization procedures, solid phase extractions (SPE) was selected as the detail characterization procedures for the micelle in order to investigate the quality attributes and material attributes of PEG-PLA micelle.
Methods: PEG-PLA were synthesized by the ring-open polymerization. Methoxy polyethyleneglycol (Mn 5kDa) was used as an initiator. DL-, L- or D- lactide were polymerized. PEG-PLA, which molecular weight of PLA domains were 5kDa and 12kDa were synthesized. Synthesized PEG-PLA were characterized by 1H-NMR, SEC-RI, DSC, CD and XRD.
PEG-PLA micelles were prepared by the anti-solvent precipitation method. PEG-PLA were dissolved in DMF, for preparing TAM loading micelle, TAM was also dissolved in DMF and then the solvent was feed in the water. Residual solvent and unloading TAM were removed by dialysis. Samples were filtrated and stored 2-8 °C until use. Prepared micelles were characterized by DLS, zeta-potentials, measuring CAC, drug loading, release rate and localizations of TAM. For investigating the localizations of TAM in the micelle, reverse phase SPE was demonstrated. Amount of TAM in the elute was determined by HPLC.
Results: Synthesized PEG-PLA was monodispersed and exerted theoretical molecular weight. The PLA domains of DL-lactide showed amorphous-like sate by DSC and XRD, on the other hand, that of L- and D- lactide showed crystalline state.
Particle size of micelle was corresponded to PLA molecular weight. Drug loading of TAM in the larger molecular weight micelles were higher than that in the smaller ones.
The amount of TAM evaluated by SPE was changed according to physicochemical properties of PEG-PLA, moreover, was corelated with the zeta-potential of the micelle loading TAM. Thereby, SPE could be detected the TAM that is adsorbed on the surface of micelle. Interestingly, PEG-PLA, which has crystalline PLA domains exerted lower adsorbed amount of TAM compared to amorphous ones. The release of TAM in the isotonic neutral buffer was also corelated with adsorbed amount of TAM.
Conclusion: It was concluded the crystallinity and molecular weight of PLA domain were critical for particle size, encapsulation efficiency and localizations in micelles and release of TAM. Therefore, they were candidates of critical material attributes. It is also important that the in vivo pharmacokinetic speculation would be achieved by in vitro characterization procedures, so we are planning to demonstrate pharmacokinetic studies and biomimetic in vitro release tests.
FDA: Food and Drug Administration, EMA: European Medicines Agency, PMDA: Pharmaceuticals and Medical Devices Agency, 1H-NMR: Proton nuclear magnetic resonance, SEC: Size exclusion chromatography, RI: Refractive index detector, DSC: Differential scanning calorimetry, XRD: X-ray diffraction, DMF: N, N-dimethylformamide, DLS: Dynamic light scattering, CAC: Critical associated concentration, HPLC: High performance liquid chromatography
Kazuyuki Takata– Toyonaka-shi, Osaka, Japan
Daiki Nomura– Amagasaki-shi, Hyogo, Japan
Yasushi Moroto– Amagasaki-shi, Hyogo, Japan
Yoshiyasu Baba– Amagasaki-shi, Hyogo, Japan
Hideyuki Kitamura– Amagasaki-shi, Hyogo, Japan