Evan Charles Unger MD, Presenter: Shareholder, NuvOx Pharma LLC Shareholder, Microvascular Therapeutics

Paul Olson DPhil, Abstract Co-Author: Employee, Kypha

Stewart Williams DPhil, Abstract Co-Author: Nothing to Disclose

Edmund Marinelli DPhil, Abstract Co-Author: Employee, NuvOx Pharma, LLC

Delphine El Mehdi PhD, Abstract Co-Author: Research, Kypha Research, PGXL Laboratories

Jeremy S Touroo BS,MS, Abstract Co-Author: Nothing to Disclose

Purpose: Uveitis is responsible for 10-20% of the cases of blindness in the U.S. and its economic impact is at least as great as diabetic retinopathy. The purpose of this research is to develop a targeted agent for molecular imaging of uveitis with ultrasound and fluorescence imaging that can be used for diagnostic purposes and also potentially as a theranostic for drug delivery.

Materials and Methods: Human retinal endothelial cells (HREC’s) were grown in 2-D static and 3-D vascular mimetic phantoms. The HRECs were exposed to lipolysaccharide (LPS) and fluorescent antibodies were used to screen for expression of E-selectin, P-selectin, ICAM-1 and VCAM. Nanoconjugates (NCs) were prepared from a blend of phospholipid (DPPC and DPPE-PEG) using mechanical agitation to entrap perfluorobutane gas. The peptide DITWDQLWDLMK-OH was incorporated into the nanoconjugates via a PEG linker attached to lipids at a mole ratio = 1%. The peptide has a 4 nM KD for murine and human E-selectin. DiI was also incorporated into the NCs. Particle sizing was performed by QELS. Imaging studies were performed with fluorescence and high frequency ultrasound (VEVO) in the 2-D and 3-D phantoms and in rats with uveitis induced by LPS (n = 5 per group).

Results: E-selectin was markedly upregulated in inflamed HREC’s, >> than the other epitopes. The mean size of the NCs was < 1-micron. In vitro imaging showed marked accumulation of NCs on inflamed HRECs in conditions of flow; NCs bearing sham peptides showed no binding. The NCs were accumulated intracellularly by the inflamed HRECs. In vivo fluorescence and US imaging showed strong signal in inflamed eyes of the rats and no uptake in controls. Ex vivo study of the rat’s eyes showed intracellular uptake of NCs by inflamed retinal endothelial cells and adjacent macrophages.

Conclusion: NCs bearing this peptide are a promising agent for molecular ultrasound and fluorescence imaging of uveitis with strong binding to both human and rat/murine epitopes of E-selectin. Intracellular uptake of NCs by inflamed endothelial cells suggests theranostic potential as a platform for drug delivery.

Because NCs work with both ultrasound and fluorescence and target a molecular marker of inflammation, NCs hold potential for better diagnosis and staging of uveitis. Intracellular uptake of NCs by inflamed cells shows potential for drug delivery.

Unger, E, Olson, P, Williams, S, Marinelli, E, El Mehdi, D, Touroo, J, Nanoconjugates as Targeted Agents for Molecular Ultrasound and Fluorescence Imaging Diagnosis and Staging of Uveitis and as Theranostic Agents.  Radiological Society of North America 2014 Scientific Assembly and Annual Meeting, - ,Chicago IL. http://archive.rsna.org/2014/14015719.html