In this study, a clear correlation between the in vitro and in vivo cellular uptake and trafficking was discovered by delivering miktoarm copolymer nanomicelles (MCNs) to cancer cells and tumor tissues. To monitor this process, two different FRET pairs, DiO and DiI, DiD and DiR, were loaded into MCNs to monitor the Förster resonance energy transfer (FRET) efficiency. The change in FRET efficiency in vitro and in vivo demonstrated a similar sequence of events for the transport of MCNs: hyperbranched block PCL inserted into cytomembrane, while the loaded hydrophobic fluorescence probes were released and followed by time-dependent intracellular clustering within endocytic vesicles. Additionally, uptake of loaded fluorescence probes with successively increasing ratios of copolymers suggested that with the increase of mass ratio of copolymer to fluorescence probes, cellular uptake of probes significantly decreased. This result was also consistent with the uptake behavior in cancer tissues. Collectively, the interaction between MCNs and cellular membrane dictated the uptake and trafficking of core-loaded hydrophobic probes. This concept paves a new way to analyze in vitro-in vivo correlation of other nanocarriers for endocytosis mechanism studies as well as further novel copolymers design in biomedical applications.
Keywords: miktoarm copolymers• nanomicelles• cellular uptake• intracellular trafficking• FRET• in vitro-in vivo correlationAdvances in nanotechnology have fostered to construct a diversity of nanomaterials with the ability to encapsulate and transport therapeutic or diagnostic agents 1-3 . Among these nanoscale delivery vehicles, polymeric micelles attracted the most attention in diverse biomedical fields as drug delivery vehicles 4-5 , contrast agents carriers 6-7 and diagnostic devices [8][9] . The advantages of polymeric micelles used as drug delivery vectors include improving the solubility of poorly water soluble drugs with the amphiphilic structures 10 , stabilizing and protecting drugs that are sensitive to the surrounding environment 11 , reducing nonspecific uptake by the reticuloendothelial system (RES) 12 and achieving the passive targeting delivery in solid tumors [13][14] . The conventional micelle drug delivery systems are mainly based on linear amphiphilic copolymers. However, linear amphiphilic copolymer micelles will dissemble in vivo when the concentration of the copolymer is diluted by the bloodstream to fall below the critical In this case, the similar chemical structures of DiO, DiI, DiD and DiR could aid us to construct dye-loaded MCNs with similar release behavior in order to discover the in vitro -in vivo correlation. (3) The FRET pair DiO and DiI, with DiO as donor and DiI as acceptor, was used to monitor FRET efficiency in vitro; while the second FRET pair DiD and DiR, with DiD as donor and DiR as acceptor was used to detect the change of FRET efficiency in vivo.