Pre‐targeted Imaging of Protease Activity through In Situ Assembly of Nanoparticles

Abstract

The pre-targeted imaging of enzyme activity has not been reported, likely owingtothe lack of amechanism to retain the injected substrate in the first step for subsequent labeling. Herein, we report the use of two bioorthogonal reactions-the condensation reaction of aromatic nitriles and aminothiols and the inverse-electron demand Diels-Alder reaction between tetrazine and trans-cyclooctene (TCO)-to develop an ovel strategy for pre-targeted imaging of the activity of proteases. The substrate probe (TCO-C-SNAT4)c an be selectively activated by an enzyme target (e.g.caspase-3/7), which triggers macrocyclization and subsequent in situ self-assembly into nanoaggregates retained at the target site.T he tetrazineimaging tag conjugate labels TCO in the nanoaggregates to generate selective signal retention for imaging in vitro,incells, and in mice.Owing to the decoupling of enzyme activation and imaging tag immobilization, TCO-C-SNAT4 can be repeatedly injected to generate and accumulate more TCO-nanoaggregates for click labeling.

Enzyme‐Mediated In Situ Self‐Assembly Promotes In Vivo Bioorthogonal Reaction for Pretargeted Multimodality Imaging

“…While TCO‐labeled NPs can be preferentially accumulated in tumors through the enhanced permeability and retention (EPR) effect, they may similarly suffer from high RES trapping and poor penetration into tumor tissues. Recently, Rao and colleagues reported an innovative pretargeting approach via combining the bioorthogonal IEDDA reaction with their developed target‐enabled in situ ligand aggregation (TESLA) strategy, allowing for fluorescence or PET imaging of caspase‐3/7 activity in therapy‐response tumors [10] . However, due to lack of an imaging moiety in the TESLA probe, it was unable to provide real‐time imaging signals to tell the optimum time for guiding subsequent injection of the Tz bearing reporters (Cy5 or 64 Cu).…”

Enzyme‐Mediated In Situ Self‐Assembly Promotes In Vivo Bioorthogonal Reaction for Pretargeted Multimodality Imaging

“…[9] While TCO-labeled NPs can be preferentially accumulated in tumors through the enhanced permeability and retention (EPR) effect, they may similarly suffer from high RES trapping and poor penetration into tumor tissues.R ecently, Rao and colleagues reported an innovative pretargeting approach via combining the bioorthogonal IEDDAr eaction with their developed target-enabled in situ ligand aggregation (TESLA) strategy,allowing for fluorescence or PET imaging of caspase-3/7 activity in therapy-response tumors. [10] However,due to lack of an imaging moiety in the TESLA probe,it was unable to provide real-time imaging signals to tell the optimum time for guiding subsequent injection of the Tz bearing reporters (Cy5 or 64 Cu). Pretargeted multimodality imaging of enzyme activity with these TESLA probes was not applicable.T ill now,arefined strategy to achieve pretargeted multimodality imaging of enzyme activity in vivo remain elusive.…”

Cancer nanotechnology: current status and perspectives

“…It is currently being studied in several clinical trials for SLN mapping, including to improve the resolution and sensitivity of nano-MRI by using a 7 Tesla scanner, particularly for small lymph nodes (ClinicalTrials.gov Identifier: NCT03280277, NCT04300673, NCT03817307, NCT02857218, NCT04261777). A precision nano-enhanced approach for monitoring disease progression utilizes triggered aggregation in the TME (Target-Enabled in situ Ligand Assembly [TESLA]) [ 301 ]. The particles are built in situ at tumor sites from precursors containing specific moieties which can form larger NPs only after being cleaved by enzymes specific to cancer cell apoptosis.…”