Multi-disciplinary study of lanthanide chelates as multi-modal molecular imaging agents

Molecular imaging is a powerful tool that has the ability to elucidate biochemical mechanisms and signal the early onset of disease. Lanthanide chelates represent a unique class of molecular imaging agents that can yield multi-modal signatures including long-lived fluorescence and magnetic resonance. The primary aim of this dissertation is to demonstrate the utility of Lanthanide chelate molecular imaging agents for contrast enhanced disease demarcation. Traditionally, Lanthanide chelate imaging agents have been non-targeted perfusion agents that distribute in-vivo based on charge and lipophilicity. It is shown here that the structural features of the chelate can be modified to facilitate tunable spectroscopic and biodistribution properties in-vivo. While our perfusion-based agents have demonstrated considerable utility, their principal limitation is specificity. For increased specificity, we synthesized a trifunctional Lanthanide chelate that possessed an antenna for metal sensitization, phosphonate acid pendant arms for chelation, and a carboxylate arm for conjugation to targeting moieties such as antibodies, peptides, or small molecule ligands.

Recently, peripheral benzodiazepine receptor (PBR) overexpression has been reported in many types of disease. Small molecule ligands of the PBR, such as PK11195, have been shown to bind with high affinity and thus could be used as contrast agent targeting moieties. Therefore, we synthesized a structural analogue of PK11195 that facilitates C-terminal conjugation. This form was then coupled to the trifunctional Lanthanide chelate and complexed with Europium and Gadolinium (Ln-PK11195).

With the Ln-PK11195 agent in hand, it was demonstrated that PBR overexpressing C6 glioma cells would actively uptake the agent and that Ln-PK11195 seemed to be localizing on the PBR. Additionally, multi-modal imaging (fluorescence and MR) was shown possible on a single group of cells incubated with a Eu-PKl 1195 and Gd- PK11195 cocktail.

Next, we demonstrated PBR profiling on surgically resected human tissue samples using Eu-PKl 1195, indicating a possible use as a histopathology stain. Both cancerous and non-cancerous PBR expressing disease was labeled with the agent.

Finally, we fully characterized the primary spectroscopic signatures of Ln- PK11195 (time-resolved fluorescence and MR) for sensitivity to pH. It was shown that Ln-PK11195 demonstrates adequate pH sensitivity to measure localized, intracellular pH in tissues.