Synthesis and Evaluation of Multimodal Near-Infrared/Technetium-99m Probes for Imaging Cancer and Bone Injury

Abstract

To address the need for better technologies for imaging disease, new strategies for the synthesis of nuclear, optical and photoacoustic (PA) multi-modal probes were developed. Initially, a library of phenyl-1H-imidazo-[4,5]-f-1,10-phenanthroline (PIP) ligands were synthesized to create isostructural optical and nuclear probes. The 99mTc complexes of the ligands were prepared in high yield and showed excellent stability in vitro. Biodistribution studies were completed where the lipophilic Tc(I) complexes had significant retention in the liver, lung, and spleen. To overcome this, a second-generation was created. Specifically, a polar cyanine dye, IR783, was linked to a PIP ligand (IR783-PIP) to reduce non-specific binding and enhance blood half-life via albumin binding. An added benefit of using IR783 is that it enabled photoacoustic (PA) imaging to be used in place of optical imaging. In vitro testing showed that it was possible to separate the PA signal of IR783-PIP from oxy- and deoxyhemoglobin. Radiolabelling of the IR783-PIP ligand with 99mTc was also achieved in 41 % yield. Because it proved difficult to target IR783-PIP, a tetrazine derived cyanine dye was prepared creating the opportunity to link the PA agent to trans-cyclooctene (TCO) modified targeting molecules. IR783-HYNIC-Tz was prepared in 2 steps in high yield through a hydrazine-based coupling strategy. The bifunctional dye bound albumin and its PA signal was separable from the signals arising in blood. In vivo studies in osteosarcoma and glioblastoma multiforme tumour models were performed where images showed clear tumour uptake. For targeting, IR783-HYNIC-Tz was used in combination with a TCO-derived bisphosphonate for imaging calcium accretion in knee and shoulder joints. The conjugate was also labelled with 99mTc which verified uptake in the joints (3-5% injected dose per gram) out to 24 hours. Lastly, the IR783-HYNIC-Tz dye was employed to tag TCO-derived gas vesicles to create a new generation of PA/ultrasound imaging agents. Overall, the strategies reported are platforms for the creation of targeted multimodal probes. They are designed to exploit biorthogonal reactions and can therefore be utilized with a wide range of targeting molecules.