Abstract
PURPOSE: The objective of this study was to formulate nanoparticles of D-luciferin (Nano-Luc), DiR (Nano-DiR) and dual functional nanoparticles with DiR and luciferin (Nano-LucDiR) for in-vivo imaging as well as tracking of the nanoparticles in tumors. METHODS: Nano-Luc and Nano-LucDiR were prepared using different lipids, and subsequently characterized for loading and entrapment efficiency, physical properties, release profile, toxicity and stability. We utilized Response Surface Methodology (RSM) to optimize the nanoparticles using design of experiment (DOE Vr.8.0). Nano-Luc was evaluated against free luciferin to establish its pharmacokinetic parameters in mice. In-vivo imaging of tumors and tracking of nanoparticles was carried out with an IVIS® Spectrum-CT (Caliper) using xenograft, orthotopic and metastatic tumor models in BALB/c nude mice with different cell lines and different routes of nanoparticle administration (subcutaneous, intraperitoneal and intravenous). RESULTS: Particle size of both Nano-Luc and Nano-LucDiR were found to be <200 nm. Nano-Luc formulation showed a slow and controlled release upto 72 h (90%) in vitro. The optimized Nano-Luc had loading efficiency of 5.0 mg/ml with 99% encapsulation efficiency. Nano-Luc and Nano-LucDiR formulations had good shelf stability. Nano-Luc and Nano-LucDiR enhanced plasma half-life of luciferin compared to free luciferin thus providing longer circulation of luciferin in plasma enabling imaging of tumors for more than 24 h. Nano-LucDiR allowed simultaneous bioluminescent and fluorescent imaging to be conducted, with three-dimensional reconstruct of tumors without losing either signal during the acquisition time. CONCLUSION: Nano-Luc and Nano-LucDiR allowed prolonged reproducible in-vivo imaging of tumors, especially during multimodality 3D imaging.