Abstract
Recent advances in the field of non-invasive optical imaging have included the development of contrast agents that report on the activity of enzymatic targets associated with disease pathology. In particular, proteases have proven to be ideal targets for development of optical sensors for cancer. Recently developed contrast agents for protease activity include both small peptides and large polymer-based quenched fluorescent substrates as well as fluorescently labeled activity based probes (ABPs). While substrates produce a fluorescent signal as a result of processing by a protease, ABPs are retained at the site of proteolysis due to formation of a permanent covalent bond with the active site catalytic residue. Both methods have potential advantages and disadvantages yet a careful comparison of substrates and ABPs has not been performed. Here we present the results of a direct comparison of commercially available protease substrates with several recently described fluorescent ABPs in a mouse model of cancer. The results demonstrate that fluorescent ABPs show more rapid and selective uptake into tumors as well as overall brighter signals compared to substrate probes. These data suggest that the lack of signal amplification for an ABP is offset by the increased kinetics of tissue uptake and prolonged retention of the probes once bound to a protease target. Furthermore, fluorescent ABPs can be used as imaging reagents with similar or better results as the commercially available protease substrates.
Highlights
The past decade has seen a dramatic increase in the number of new technologies that are available for applications in molecular imaging and disease monitoring
Recent advances in the development of new classes of optical sensors have had a profound impact on the field of non-invasive fluorescence imaging
Perhaps one of the most promising new approaches for the development of optical contrast agents is the development of ââsmart probesââ that produce a specific signal as the result of the action of a given enzymatic target
Summary
The past decade has seen a dramatic increase in the number of new technologies that are available for applications in molecular imaging and disease monitoring. At the heart of all new imaging methods is the need for contrast agents that provide a more precise picture of distinct molecular events as they happen in vivo. Because increased protease activity has been shown to be associated with the pathogenesis of a number of human diseases including cancer, atherosclerosis and neurodegenerative diseases, significant efforts have been made to develop molecular sensors of protease activity. A large number of reagents have been built around reporter substrates that, when cleaved by a protease, produce a fluorescent signal (for review see[1,2]). These reagents include large, polymer-based quenched fluorescent substrates that are cleaved in multiple locations to produce fluorescent products (Fig. 1A). As an alternative to substrates, fluorescent activity based probes have been reported [3,4]
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