Systematic evaluation and hepatic targeting of a dual-fluorescence icaritin microemulsion: elucidating the protein corona-mediated delivery mechanism.

Tracking the in vivo fate of nanodrugs remains a significant challenge. In this study, we developed a dual-fluorescence-labeled macromolecular lipid (TPE-PEG2000-ICG; TIP) and a corresponding dual-fluorescence-labeled icaritin microemulsion (TIP-IC-MEs) that could in real time trace both nanocarrier distribution and structural integrity. The design couples the aggregation-caused quenching (ACQ) of indocyanine green (ICG) with the aggregation-induced emission (AIE) of tetraphenylethylene (TPE) via a flexible polyethyleneglycol 2000 (PEG2000) chain. The TPE-AIE moiety could enhance the ACQ effect of the ICG channel, greatly increasing its quenching sensitivity in aqueous environments. This improvement significantly reduced the fluorescence recovery rate of TIP upon probe release from the formulation, which improved the accuracy and reliability of TIP when studying the in vivo fate of nanoformulations. TIP-IC-MEs exhibited favorable physicochemical stability, pH-responsive drug release and in vivo safety. In vitro and in vivo studies further elucidated the critical role of the protein corona in modulating hepatic targeting and clearance pathways of TIP-IC-MEs. Proteomic analysis demonstrated that TIP-IC-MEs selectively recruited specific protein corona components associated with immune recognition, cellular transport, and metabolic reprogramming within the hepatocellular carcinoma model. These corona constituents collectively facilitated tumor-targeted delivery of TIP-IC-MEs through a "cell-hitchhiking" mechanism. This study revealed a dual, context-dependent regulatory role of the protein corona on the in vivo behavior of nanoformulations, providing a novel imaging technology and a mechanistic foundation for rational nanocarrier design and supports the clinical translation of nanomedicines.