Development of a protease-sensitive molecular imaging agent for optoacoustic tomography

We are working to develop a molecular imaging agent that will allow for in vivo imaging of proteases by use of optoacoustic tomography. Proteases are protein-cleaving proteins known to be overactive in a number of pathologies, including cancers and vascular disease. Protease-sensitive "smart probes" have previously been developed in the context of pure optical imaging. These involve pairs of mutually quenching fluorophores attached to a backbone by protease-cleavable peptide side chains; cleaving of the side chains liberates the fluorophores and leads to increase in fluorescence. Optoacoustic imaging is sensitive not to fluorescence but to optical absorption and so a smart imaging probe for protease imaging would need to shift its absorption peak upon cleavage. Naturally, the absorption peaks of the cleaved (and, ideally, uncleaved) molecules should be in the near infrared for maximum tissue penetration. We have designed a molecule that should achieve these specifications. It comprises two active sites, derivatives of natural photosynthetic bacteriochlorophylls that absorb in the near IR, conjugated to a lysine backbone by peptide spacers specific to the protease being imaged. When these bacteriochlorophylls dimerize and stack in the uncleaved molecule, their absorption peak shifts about 20-30 nm. When they are cleaved from the molecule the absorption peak shifts back to that of bacteriochlorophyll monomers. We have performed a preliminary synthesis of the molecule and confirmed by use of a spectrometer that the pairing of the bacteriochlorophylls leads to the expected absorption shift.