Activatable chemiluminescence probe based on four-arm PEG-conjugated-pyropheophorbide-a for in vivo autofluorescence-free imaging of peroxynitrite

Abstract

Peroxynitrite (ONOO⁻) is a highly reactive nitrogen species that plays pivotal roles in cell signal transduction and physiological or pathological progresses. However, commonly used ONOO⁻ optical imaging probes are still hampered by high background/autofluorescence (fluorescence probe), short emission wavelength, or poor selectivity in the case of chemiluminescence. Herein, we report a facile method to prepare an activatable chemiluminescence probe (PPA-PEG) with good biocompatibility and functionality for in vivo autofluorescence-free imaging of ONOO⁻. The PPA-PEG consists of pyropheophorbide-a (PPA), a typical deep red photosensitizer that acts as both the recognition and signaling element, and 4-arm poly(ethylene glycol) (4-arm PEG), which improves the biosafety and water solubility of probe. These components can self-assemble into nanoparticles (namely PPA-PEG nanoprobe) in aqueous solution. The PPA-PEG nanoprobe showed an ultra-low chemiluminescence signal before interacting with ONOO⁻, but exhibited good selectivity, high sensitivity and a fast response toward ONOO⁻. The PPA-PEG was successfully applied to image cellular ONOO⁻ changes, as well as the endogenous ONOO⁻ changes in inflammation models and subcutaneous or orthotopic hepatocellular carcinoma (HCC) tumors models in living mice. In vitro and in vivo studies verified the good detection and imaging capabilities of PPA-PEG for peroxynitrite, demonstrating suitable tissue penetration and a high signal-to-background ratio (SBR). Thus, our nanoprobe can serve as a valuable activatable chemiluminescence imaging tool for studying important peroxynitrite-related chemical and biological applications.