Analysis of Stable Chelate-free Gadolinium Loaded Titanium Dioxide Nanoparticles for MRI-Guided Radionuclide Stimulated Cancer Treatment

Background: Recent studies demonstrate that titanium dioxide nanoparticles (TiO2 NPs) are an effective source of reactive oxygen species (ROS) for photodynamic therapy and radionuclide stimulated dynamic therapy (RaST). Unfortunately, tracking the in vivo distribution of TiO2 NPs noninvasively remains elusive. Objective: Given the use of gadolinium (Gd) chelates as effective contrast agents for magnetic resonance imaging (MRI), this study aims to (1) develop hybrid TiO2-Gd NPs that exhibit high relaxivity for tracking the NPs without loss of ROS generating capacity; and (2) establish a simple colorimetric assay for quantifying Gd loading and stability. Method: A chelate-free, heat-induced method was used to load Gd onto TiO2 NPs, which was coated with transferrin (Tf). A sensitive colorimetric assay and inductively coupled plasma mass spectrometry (ICP-MS) were used to determine Gd loading and stability of the TiO2-Gd-Tf NPs. Measurement of the relaxivity was performed on a 1.4 T relaxometer and a 4.7 T small animal magnetic resonance scanner to estimate the effects of magnetic field strength. ROS was quantified by activated dichlorodihydrofluorescein diacetate fluorescence. Cell uptake of the NPs and RaST were monitored by fluorescence microscopy. Both 3 T and 4.7 T scanners were used to image the in vivo distribution of intravenously injected NPs in tumor-bearing mice. Results: A simple colorimetric assay accurately determined both the loading and stability of the NPs compared with the expensive and complex ICP-MS method. Coating of the TiO2-Gd NPs with Tf stabilized the nanoconstruct and minimized aggregation. The TiO2-Gd-Tf maintained ROS-generating capability without inducing cell death at a wide range of concentrations but induced significant cell death under RaST conditions in the presence of F-18 radiolabeled 2-fluorodeoxyglucose. The longitudinal (r1 = 10.43 mM^-1s^-1) and transverse (r2 = 13.43 mM^-1s^-1) relaxivity of TiO2-Gd-Tf NPs were about twice and thrice, respectively, those of clinically used Gd contrast agent (Gd-DTPA; r1 = 3.77 mM^-1s^-1 and r2 = 5.51 mM^-1s^-1) at 1.4 T. While the r1 (8.13 mM^-1s^-1) reduced to about twice that of Gd-DTPA (4.89 mM^-1s^-1) at 4.7 T, the corresponding r2 (87.15 mM^-1s^-1) increased by a factor 22.6 compared to Gd-DTPA (r2 = 3.85). MRI of tumor-bearing mice injected with TiO2-Gd-Tf NPs tracked the NPs distribution and accumulation in tumors. Conclusion: This work demonstrates that Arsenazo III colorimetric assay can substitute ICP-MS for determining the loading and stability of Gd-doped TiO2 NPs. The new nanoconstruct enabled RaST effect in cells, exhibited high relaxivity, and enhanced MRI contrast in tumors in vivo, paving the way for in vivo MRI-guided RaST.