Imaging of Blood-Brain Barrier Disruption by Tc-99m-Tilmanocept Molecular Imaging of a Rodent Model of Traumatic Brain Injury

276 Objectives: The blood-brain barrier (BBB) disruption and subsequent neuro-inflammation have been implicated in a spectrum of human disorders and injuries. Some examples are: traumatic brain injury, stroke, multiple sclerosis, and Alzheimer’s Disease. Technetium-99m-labeled tilmanocept is a 18 kDa molecule with a hydrodynamic radius of 3.5 nm designed for selective molecular imaging of tissues bearing the CD206 receptor. Initially proposed for sentinel lymph node mapping, the radiopharmaceutical was approved by the U.S. FDA in 2013 and the EMA in 2014. CD206 has been isolated from the surface membrane of perivascular microglial cells. We hypothesis that microglial cells bearing CD206 will trap Tc-99m-tilmanocept after migration through a disrupted blood-brain barrier. Methods: Eight male age-matched mice were used in this study: four served as healthy controls and four received a controlled cortical impact. The latter four mice were anesthetized and fixed into a stereotactic frame. A burr hole was made approximately 5 mm anterior to posterior (10 to -45 A-P) from the bregmatic suture and 4 mm laterally from the sagittal suture over the right hemisphere. Craniotomies were made with a portable drill over the right parietotemporal cortex and the bone flap was removed. Using a stereotaxic impactor, a 3-mm tip was accelerated down to a 1.5 mm depth at a speed of 3.0 m/sec with a 45 msec dwell time. Five days following the brain injury the eight mice were injected (i.p.) with FITC-HSA. Two hours later each animal was anesthetized, injected (i.v., 0.015 nmol/g, 2 mCi) with IRDye800CW-labeled Tc-99m-tilmanocept, positioned atop a high-resolution gamma camera, and imaged. At 45 mins post injection, the mice were perfused with 4% zinc-formalin solution and their brain removed to assay the amount of accumulated radioactivity and to detect BBB disruption (FITC-HSA) and IRDye800CW-labeled tilmanocept tissue distribution via fluorescence imaging of brain cross-sections. Results: The Tc-99m-tilmanocept accumulation by the TBI brains (0.050 ±0.013 %/g) was higher (P = 0.001) than the healthy brains (0.020 ±0.002 %/g). Because the brains were perfused prior to excision, the uptake represents receptor-bound Tc-99m-tilmanocept only, and does not represent unbound tilmanocept within the blood. Most importantly, the amount of radioactivity within each of the TBI brains was higher than the mean activity of the healthy brains by greater than 3 times the standard deviation of the healthy brain activities. The %ID/g measurement were consistent with 30-45 min PI static images, which displayed higher count rates by the TBI mice. Near infra-red imaging (NIR) of brain histo-micrographs demonstrated co-localization of fluorescent-tilmanocept with FITC-HSA. NIR or FITC signals could not be found in the brain cross-sections of the healthy mice. Conclusions: These results are consistent with the hypothesis that microglial cells bearing CD206 trap fluorescent-labeled Tc-99m-tilmanocept in regions of BBB disruption. This provides the motivation for a clinical trial of intra-venously administered Tc-99m-tilmanocept with the goal of developing a SPECT/CT protocol for the detection of neuro-inflammation.