Radioactive smart probe for corrected matrix metalloproteinase imaging

Objectives Although various activatable optical probes have been developed to visualize the metalloproteinases (MMPs) activities in vivo, precise quantification of the enzyme activity is limited due to the inherent scattering and attenuation (limited depth penetration) properties of optical imaging. We are developing a series of nontargeted and targeted radioactive smart probes for corrected matrix metalloproteinase imaging.

Methods The peptides containing the MMP sensitive sequence were prepared through Fmoc Solid Phase Peptide Synthesis (SPPS). After conjugation with fluorescent dyes BBQ650 and Cy5.5, a novel activatable peptide probe ⁶⁴Cu-BBQ650^TM-(MMP)-K(Cy5.5)-E-K(DOTA)-OH was constructed to detect the tumor MMP activity. In vitro and proof-of-principle in vivo experiments were performed to study the probe’s MMP sensitivity.

Results The agent ⁶⁴Cu-BBQ650^TM-(MMP)-K(Cy5.5)-E-K(DOTA)-OH is optically quenched in its native form, but demonstrated an 8.2-fold increase in fluorescence upon the cleavage by MMP-13 enzyme. The in vivo MMP activation was confirmed in U87MG tumor model, which could be successfully blocked with MMP inhibitor. Clearly, introducing the PET motif did not significantly change the peptide’s MMP sensitivity. We demonstrated that ⁶⁴Cu-BBQ650^TM-(MMP)-K(Cy5.5)-E-K(DOTA)-OH could clearly reflect the fivefold concentration difference of the localized probes (with microPET), which could be used to correct the enzyme activity determined from optical imaging.

Conclusions The use of a single modality to image biomarkers/processes may limit detailed information regarding tracer localization and targeted enzyme activity. A coincident measurement technique coupled with optical imaging may be advantageous, as we have demonstrated the feasibility of correcting the activatable optical imaging data through the PET signal. This approach stands as an attractive new strategy for accurate imaging of MMP activities, which can also be applied for other protease imaging.

Research Support This work was supported by the USC Department of Radiology, the American Cancer Society (for Dr. Li, 121991-MRSG-12-034-01-CCE), and the USC Biomedical Imaging Science Initiative