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Remote loading of liposomes by transmembrane gradients is one of the best approaches for achieving the high drug level per liposome. This breakthrough, which enabled the clinical use of nanoliposomal drugs, has not been paralleled by good understanding that allows predicting loading efficiency of drugs. We developed a model that relates drug physicochemical properties and loading conditions to loading efficiency. This model enables choosing candidate molecules for remote loading and optimizing loading conditions according to logical considerations. In order to improve the therapeutic efficacy of two drug combination, vincristine and topotecan were co- remote loaded by ammonium sulfate gradient in the same nanoliposome (LipoViTo). The nanoliposomes controlled the drugs´ biofate and maintained a fixed drug ratio in vivo. Pharmacokinetics studies showed that LipoViTo deliver the two drugs simultaneously to the tumors, where they are released at a predefined ratio. LipoViTo were more efficacious than the free drugs and liposomes withone agent, singly or in combination. Our approach should improve the development of liposomal formulations for clinical applications.

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