Response to Comment on “A Liposomal System Capable of Generating CO₂ Bubbles to Induce Transient Cavitation, Lysosomal Rupturing and Cell Necrosis”

Abstract

ammonium bicarbonate ·carbon dioxide ·cavitation · drug delivery ·liposomes Leroux et al. performed aset of experiments reported in our study and claimed in aC orrespondence in this journal that they failed to reproduce the thermoresponsive liposomal system containing ammonium bicarbonate (ABC) that could trigger drug release under mild heating. [14] We disagree with their assessments and speculate that the different method used by Leroux et al. to prepare the aqueous ABC is the culprit. Leroux et al. reported ac onstant generation of CO 2 bubbles during the preparation of their ABC liposomes;w e observed no such phenomenon. As tudy of recent literature indicates that nine independent groups have been able to duplicate the findings described in our original publication. We stand by the core findings of our study.Ammonium bicarbonate (ABC;NH 4 HCO 3 )isoften used as araising agent in baking;upon heating, ABC decomposes to generate CO 2 bubbles.I naCommunication published in this journal in 2012, [1] we proposed and demonstrated that this unique compound could be encapsulated in the interiors of liposomes to destruct the lipid bilayer membranes and thereby trigger drug release upon mild heating. To demonstrate this concept, we loaded both ABC and calcein into the aqueous cores of liposomes using the conventional lipid-film hydration technique.F igure 1a,b presents results concerning the production of CO 2 bubbles from the ABC liposomes under mild heating at 42 8 8Cand their calcein release profiles. Thel ipid-film hydration technique is generally regarded as not an ideal method for loading an active drug into liposomes. In our subsequently published papers,the lipid-film hydration technique was used to prepare the ABC liposomes and then ar emote-loading technique was used to achieve highly efficient doxorubicin encapsulation, exploiting the transmembrane gradient of NH 4 + ,i namanner similar to that in the production of conventional ammonium-sulfate liposomes.We disagree with Leroux et al. that ABC liposomes cannot be prepared as as table system because of the transmembrane osmotic gradient that is established by the loaded ionic species.When the lipid-film hydration technique is used to prepare liposomes,t he transmembrane osmotic gradient is negligible if an intact system can be obtained, as the media inside and outside of intact liposomes are similar. During the purification process,t he as-prepared liposomes