A Targeting Microbubble for Ultrasound Molecular Imaging.

James Shue-Min Yeh,Dorian O Haskard,Sussan Nourshargh,Ellen Mcconnell,John M Seddon,Meng-Xing Tang,Charles A Sennoga,Robert Eckersley,Petros Nihoyannopoulos,Christoph E Hagemeyer

doi:10.1371/journal.pone.0129681

Abstract

RationaleMicrobubbles conjugated with targeting ligands are used as contrast agents for ultrasound molecular imaging. However, they often contain immunogenic (strept)avidin, which impedes application in humans. Although targeting bubbles not employing the biotin-(strept)avidin conjugation chemistry have been explored, only a few reached the stage of ultrasound imaging in vivo, none were reported/evaluated to show all three of the following properties desired for clinical applications: (i) low degree of non-specific bubble retention in more than one non-reticuloendothelial tissue; (ii) effective for real-time imaging; and (iii) effective for acoustic quantification of molecular targets to a high degree of quantification. Furthermore, disclosures of the compositions and methodologies enabling reproduction of the bubbles are often withheld.ObjectiveTo develop and evaluate a targeting microbubble based on maleimide-thiol conjugation chemistry for ultrasound molecular imaging.Methods and ResultsMicrobubbles with a previously unreported generic (non-targeting components) composition were grafted with anti-E-selectin F(ab’)2 using maleimide-thiol conjugation, to produce E-selectin targeting microbubbles. The resulting targeting bubbles showed high specificity to E-selectin in vitro and in vivo. Non-specific bubble retention was minimal in at least three non-reticuloendothelial tissues with inflammation (mouse heart, kidneys, cremaster). The bubbles were effective for real-time ultrasound imaging of E-selectin expression in the inflamed mouse heart and kidneys, using a clinical ultrasound scanner. The acoustic signal intensity of the targeted bubbles retained in the heart correlated strongly with the level of E-selectin expression (|r|≥0.8), demonstrating a high degree of non-invasive molecular quantification.ConclusionsTargeting microbubbles for ultrasound molecular imaging, based on maleimide-thiol conjugation chemistry and the generic composition described, may possess properties (i)–(iii) desired for clinical applications.

Highlights

Ultrasound molecular imaging has been achieved using targeting microbubbles which contain targeting ligands on the bubble-shell [1]
Targeting microbubbles for ultrasound molecular imaging, based on maleimide-thiol conjugation chemistry and the generic composition described, may possess properties (i)–(iii) desired for clinical applications
About half of them were tested in animals for ultrasound molecular imaging (Table A in S1 File), none were reported/evaluated to show all three of the following properties desired for clinical applications: (i) low degree of non-specific bubble retention in more than one non-reticuloendothelial tissue; (ii) effective for real-time imaging (allowing ready visualization of the molecular target(s) by the bedside, and better assessment of moving objects such as the beating heart); and (iii) effective for quantification of molecular targets to a high degree of quantification

Summary

Introduction

Ultrasound molecular imaging has been achieved using targeting microbubbles which contain targeting ligands on the bubble-shell [1]. Following intravenous (iv) administration, these echogenic bubbles circulate and accumulate in regions expressing the molecules targeted, depicted on ultrasound pictures as areas of bright signals locating the molecules of interest. This technique has allowed molecular imaging of pathophysiological processes such as inflammation, angiogenesis and thrombosis, and has potential for clinical applications (Table A in S1 File). Targeting bubbles often employed biotin-(strept)avidin conjugation chemistry for grafting targeting ligands to the bubble-shell (Table A in S1 File), which is best avoided for use in humans because of the high immunogenicity of (strept)avidin. Enabling disclosure of the compositions and methodologies for reproducing these bubbles are often withheld (Table A in S1 File)

Methods

Results

Conclusion