Cell-Permeable, MMP-2 Activatable, Nickel Ferrite and His-tagged Fusion Protein Self-Assembled Fluorescent Nanoprobe for Tumor Magnetic Targeting and Imaging.

Matrix metalloproteinases (MMP) activatagble imaging probe has been explored for tumor detection. However, activation of the probe is mainly occurred in the extracellular space without intracellular uptake of the probe for more sensitivity. Whilst cell penetrating peptides (CPPs) have been demonstrated to enable intracellular delivery of the imaging probe, they nevertheless encounter off-target delivery of the cargos to normal tissues. Herein, we developed a dual MMP-2-activatable and tumor cell-permeable magnetic nano-probe to simultaneously achieve selective and intracellular tumor imaging. This novel imaging probe was constructed by self-assembling of a His-tagged fluorescent fusion protein chimera and nickel ferrite nanoparticles via a chelation mechanism. The His-tagged fluorescent protein chimera consisted of a red fluorescent protein mCherry that acted as the fluorophore, the LMWP (Low Molecular Weight Protamine) peptide as a CPP, and the MMP-2 cleavage sequence fused with the hexa-histidine tag, whereas the nickel ferrite nanoparticles functioned as the His-tagged protein binder and also the fluorescent quencher. Both in vitro and in vivo results revealed that this imaging probe would not only remain non-permeable to normal tissues thereby offsetting the non-selective cellular uptake but was also suppressed of fluorescent signals during magnetic tumor targeting in the circulation, primarily due to masking of the CPP activity and quenching of the fluorophore, respectively, by the associated NiFe2O4 nanoparticles. However, these properties were recovered or "turned on" by the action of tumor-associated MMP-2 stimuli, leading to cell penetration of the nano-probes as well as fluorescence restoration and visualization within the tumor cells. To this regard, the presented tumor-activatable and cell-permeable system deems to be an appealing platform to achieve selective tumor imaging and intracellular protein delivery. Its impact is therefore significant, far-reaching and wide-spread.