Combining Doxorubicin-Loaded PEGylated Poly(Lactide-co-glycolide) Nanoparticles with Checkpoint Inhibition Safely Enhances Therapeutic Efficacy in a Melanoma Model.

Doxorubicin (DOX) has been widely used for the treatment of various cancers, however, the use of soluble DOX is limited by its low therapeutic index and improved formulations are therefore sought. Aside from its tumoricidal properties, DOX has also been shown to cause an immunogenic form of cell death, however, it is becoming abundantly clear that in situ immune stimulation alone is insufficient to cause significant immune based antitumor activity and that immune checkpoint modulation is also required. In this study, DOX-loaded nanoparticles were made by nanoprecipitation of DOX with a PEGylated poly(lactide--glycolide) copolymer (DOX-PLGA-PEG NPs) and were then tested in combination with immune checkpoint blockade (antiprogrammed death (anti-PD-1)) in a murine melanoma model to enhance antitumor effectiveness. Results showed the prepared particles to be approximately 134 nm in diameter (zeta potential -22 mV) with a loading of 1.75 μg DOX/mg NPs. In vitro release studies (of DOX) revealed the NPs to exhibit a 12 h burst release phase, followed by a slower release phase for up to 200 h. Survival studies of mice challenged with B16.F10 melanoma cells, revealed 60% of mice treated with the combination of DOX-PLGA-PEG NPs plus anti-PD-1 were tumor-free at the completion of the study. This combination therapy demonstrated higher antitumor efficacy in vivo compared to control, soluble DOX, and monotherapy of DOX-PLGA-PEG NPs or anti-PD-1 solution ( < 0.05). Moreover, in vivo safety studies (mouse weight/histopathological/toxicity) were investigated and results suggested that the combination therapy was safe. In conclusion, this study demonstrates the successful fabrication of DOX-loaded NPs by a nanoprecipitation method, and when combined with checkpoint inhibition could provide significant therapy in a murine melanoma model, suggesting that the DOX-PLGA-PEG NPs may be generating immune stimulation in situ and that benefit from this combination may be obtained in a clinical setting in the future.