Efficient tumor regression by a single and low dose treatment with a novel and enhanced formulation of thermosensitive liposomal doxorubicin.

We have developed a novel and simplified thermosensitive liposomal formulation (HaT: Hyperthermia-activated cytoToxic) composed of DPPC lipid and Brij78 (96:4, molar ratio). The HaT nanoparticles were loaded with doxorubicin (DOX) with >95% efficiency when a pH gradient method and a drug/lipid ratio of 1/20 (w/w) were applied. Drug release from the HaT formulation was significantly faster at 40-41°C (100% release in 2-3min) with 3.4-fold increased membrane permeability compared to the LTSL (lyso-lipid temperature sensitive liposomes; DPPC: MSPC: DSPE-PEG(2000)=86:10:4, molar ratio), a formulation that is currently in clinical trials. Both formulations displayed similar stability at 37°C in serum (10-20% release in 30min), which corresponds to their comparable pharmacokinetics in the unheated mice. An approximately 1.4-fold increased drug delivery to the locally heated tumor (~43°C) was detected with HaT-DOX compared to LTSL-DOX. Moreover, when compared with free DOX, HaT enhanced drug uptake in the heated tumor by 5.2-fold and reduced drug delivery to the heart by 15-fold. A single i.v. treatment with HaT-DOX at 3mg DOX/kg in combination with localized hyperthermia demonstrated enhanced tumor regression compared to LTSL-DOX and free DOX, and exhibited little toxicity.