A novel method to load topotecan into liposomes driven by a transmembrane NH₄EDTA gradient

Antitumor drugs not only cause cytocidal effect on cancer cells, but also damage on normal healthy tissues, resulting in side effects. Liposome encapsulation can result in reduced systematic distribution due to the enhanced permeability and retention (EPR) effect, accompanied by drug accumulation in liver, spleen, and other immune organs, which can cause damage to those organs. It has been demonstrated that EDTA, frequently used as a chelator, possesses a synergistic antitumor effect. Indeed, our previous study showed that EDTA could reduce the toxicity of anthracyclines to the heart and immune organs. In this study, we intended to encapsulate topotecan within liposome adopting transmembrane NH(4)EDTA gradient in order to increase the antitumor activity and decrease the toxicity against normal immune organs. Regarding the encapsulation efficiency of topotecan liposomes, both the pH value of the buffer and the cholesterol content showed significant effects on encapsulation and drug retention. Liposome encapsulation dramatically increased the antitumor activity of topotecan compared to free drug (p<0.05), while similar efficacy was obtained from liposomes prepared by a NH(4)EDTA gradient or a (NH(4))(2)SO(4) gradient (tumor inhibition ratios were 85.6% and 84.1%, respectively). However, a significant decrease in toxicity against the immune organs was found in liposomes prepared by a NH(4)EDTA gradient compared to those prepared by a (NH(4))(2)SO(4) gradient. These results suggest the superiority of the proposed gradient for topotecan encapsulation in decreasing its toxicity on immune systems.