Encapsulation of doxorubicin into thermosensitive liposomes via complexation with the transition metal manganese

In the present study, doxorubicin was encapsulated into two thermosensitive liposome formulations which were composed of DPPC / MSPC / DSPE-PEG 2000 (90 / 10 / 4 mole ratio) or DPPC/DSPE-PEG 2000 (95/5 mole ratio). Doxorubicin loading was achieved through the use of a pH gradient or a novel procedure that involved doxorubicin complexation with manganese. Regardless of the initial drug-to-lipid ratios (D : L), the final D : L reached a maximum of 0.05 (w / w) when doxorubicin was encapsulated via a pH gradient for both thermosensitive liposome formulations. In contrast, the final maximum D : L achieved through manganese complexation was 0.2 (w / w), and this loading method did not affect temperature-induced drug release, with 85% of drug released from MSPC-containing liposomes within 10 min at 42 °C but < 5% released over 60 min at 37 °C. When the thermosensitive liposomes prepared via the two different loading methods were injected into mice, similar plasma elimination profiles were observed. Cryo-transmission electron microscopy analysis indicated the presence of doxorubicin fiber bundles in liposomes loaded via pH gradient, compared to a stippled and diffuse morphology in those loaded via manganese complexation. To investigate the effect of intraliposomal pH on drug precipitate morphology, the A23187 ionophore (mediates Mn 2+/H + exchange) was added to liposomes loaded with doxorubicin–manganese complex, and the stippled and diffuse appearance could be converted to one exhibiting fiber bundles after acidification of the liposome core. This suggests that the formation of doxorubicin–manganese complex is favored when the intraliposomal pH is > 6.5. During the conversion to the fiber bundle morphology, no doxorubicin release was observed when A23187 was added to liposomes exhibiting a 0.05 (w / w), whereas a significant release was noted when the initial D : L was 0.2 (w / w). Following acidification of the liposomal interior and establishment of an apparent new D : L equilibrium, the measured D : L ratio was 0.05 (w / w). In conclusion, the manganese complexation loading method increased the encapsulation efficiency of doxorubicin in thermosensitive liposomes with no major impact on temperature-triggered drug release or pharmacokinetics.