Highly selective in-vivo imaging of tumor as an inflammation site by ROS detection using hydrocyanine-conjugated, functional nano-carriers

Abstract

Previously, the optical imaging of chitosan-functionalized, Pluronic-based nano-carriers by Cy5.5 conjugation revealed a good tumor targeting characteristic of the nano-carriers in vivo [J. Control. Release, 147 (2010) 109–117]. However, in spite of the relatively strong signal from tumor site, they also showed strong fluorescence signals from other organs, especially liver. Thus, for the detection of pathological sites, the direct use of the Cy5.5-conjugated nano-carriers is limited due to significant background signals associated with non-specific delivery of the probes. To overcome this limitation, in this study, we prepared hydrocyanine-conjugated and chitosan-functionalized Pluronic-based nano-carriers (Hydrocyanine-NC) that can detect ROS in pathological sites. The reduction of cyanine to hydrocyanine of the nano-carriers resulted in complete disappearance of fluorescence emission, and the fluorescence could be recovered by ROS-induced re-oxidization. Hydrocyanine-NC could detect various ROS including superoxide anion (O 2 −) and hydroxyl radical (OH −) in a dose-dependent manner. Hydrocyanine-NC was also stable in serum-containing media and did not show acute cytotoxicity. Hydrocyanine-NC developed strong fluorescence by the intracellular ROS formation in LPS-stimulated macrophage cells in vitro. As an in-vivo inflammation site imaging, SCC7 tumor-bearing mice were optically monitored after the i.v. injection of the dye-conjugated nano-carriers. When non-reduced, cyanine-conjugated and chitosan-functionalized Pluronic-based nano-carriers (Cyanine-NC) were injected, strong fluorescence emission was observed from the abdominal area as well as from the tumor site, and it remained over 2 days. In contrast, in the case of Hydrocyanine-NC, the initially very weak fluorescence emission from the abdominal area disappeared over time whereas the fluorescence emission from the tumor site was similar to that of Cyanine-NC. Therefore, the re-oxidation of Hydrocyanine-NC by ROS in vivo specifically eliminated the background signals from non-specific delivery of the probes, but it produced fluorescence emission strong enough to monitor the target inflammation site selectively.