Molecular imaging techniques have demonstrated efficacy in the setting of preclinical research, primarily for drug discovery and basic science. Optical imaging has emerged as a market leader in the field, owing to the availability of easy-to-use contrast agents and cost-effective imaging equipment. Recently, near-infrared (NIRF) imaging has seen the development of numerous highly sensitive contrast agents, which generally consist of a fluorophore coupled to a peptide, antibody, or nanoparticle. Two potential drawbacks to optical imaging are related to the target-to-background ratio of the contrast agents, and the time required for clearance of the contrast. We propose to investigate a novel micro particle contrast agent for optical imaging of inflammation. Targeson has developed a gas-encapsulated micro bubble that is currently used as a contrast agent for ultrasound imaging. The micro bubbles have a mean diameter of 2.5 um, and ligands (antibodies, peptides, and glyococonjugates) can be readily coupled to the surface of the micro bubble for the purpose of molecular imaging. The micro bubble contrast agent is useful for imaging molecular targets expressed on the luminal vascular endothelium;micro bubbles targeted to various molecules of relevance to inflammation and angiogenesis have demonstrated efficacy in mouse studies. The target specificity of the micro bubble is in part due to the confinement of the micro bubble to the intravascular space, rendering it a purely intravascular tracer. Micro bubbles that are not retained at the target site are cleared to the liver, spleen and lungs within minutes after administration, providing an exceptionally low background contrast signal. Additionally, micro bubbles can be readily destroyed by high-intensity ultrasound. This results in dissolution of the gas core and fragmentation of the lipid shell. In the context of ultrasound molecular imaging, this property enables users to administer sequential contrast agents in the same imaging setting;agents within the target tissue are cleared between scans by insonation of the tissue. We hypothesize that a micro bubble bearing a NIRF reporter could be an efficacious contrast agent for optical imaging of endothelial molecular targets. The large size of the micro bubble relative to single-molecule contrast agents enables a significantly larger payload of fluorophore, and may yield greater specificity owing to the intravascular confinement of the micro bubble. Additionally, the ability to clear the micro bubbles from the target tissue may enable high-throughput optical imaging of multiple molecular targets within the same imaging setting.
We aim to derivatize our micro bubble platform with an antibody against the pro-inflammatory molecular target P-selectin, and conjugate a NIRF reporter 1) on the surface and 2) within the shell of the micro bubble. We will assess the ability of these micro bubbles to bind recombinant P-selectin in vitro and in vivo by intravital microscopy. Finally, we will assess the efficacy of these agents to detect P-selectin expression in a mouse model of inflammation using NIRF whole body optical imaging. Successful completion of this project will result in a contrast agent for optical imaging that could enable a significant increase in throughput, and be readily marketed to existing optical imaging users.
Probes able to detect molecular components have the potential to accelerate drug discovery, enhance basic research, and improve the sensitivity of disease detection in a clinical setting. Optical imaging is widely used in research applications, and provides a cost-effective and reproducible imaging technique that does not require ionizing radiation. The optical imaging contrast agents described here will enable a substantial increase in the throughput and versatility of optical imaging in the preclinical research setting.
