Abstract
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Objectives To design, synthesize, and characterize multifunctional unimolecular micelles, made of hyperbranched amphiphilic block copolymers, for cancer-targeted drug delivery and non-invasive positron emission tomography (PET) imaging in tumor-bearing mice.
Methods The hyperbranched amphiphilic block copolymer, Boltorn® H40-poly(L-glutamate-hydrazone-doxorubicin)-b-poly(ethylene glycol), was conjugated with cyclic RGD peptides (cRGD, for integrin αvβ3 targeting) and macrocyclic chelators (NOTA, for 64Cu-labeling). The final conjugate was named as "H40-DOX-cRGD", where DOX (i.e. doxorubicin) was covalently linked via a pH-labile hydrazone linkage to enable pH-controlled drug release. H40-DOX (without cRGD) was also synthesized as a control. Flow cytometry, confocal microscopy, and MTT assay were carried out to measure the integrin αvβ3 targeting characteristics of H40-DOX-cRGD in vitro. PET imaging, biodistribution and blocking studies were performed to evaluate the tumor targeting efficacy of H40-DOX-cRGD in vivo, which was further validated by ex vivo experiments.
Results The unimolecular micelles exhibited a uniform size distribution (36±3 nm) and pH-sensitive drug release behavior.H40-DOX-cRGD had much higher cellular uptake in U87MG cancer cells (due to integrin αvβ3-mediated endocytosis) than H40-DOX, which led to higher cytotoxicity. In U87MG tumor-bearing mice, 64Cu-labeled H40-DOX-cRGD also exhibited significantly higher tumor uptake (4.1±0.5, 5.7±1.2, 3.7±0.8, and 3.1±0.2 %ID/g at 0.5, 4, 16, and 24 h p.i. respectively) than H40-DOX-64Cu , which was confirmed by biodistribution studies. Furthermore, ex vivo optical imaging of DOX fluorescence confirmed that DOX was indeed delivered to the tumor at a much higher level by H40-DOX-cRGD (i.e. active targeting) than H40-DOX (i.e. passive targeting).
Conclusions These unimolecular micelles that synergistically integrated passive and active tumor-targeting abilities, pH-controlled drug release, PET imaging capabilities, and suitable size range, are promising drug nanocarriers for cancer theranostics