A genetic reporter for visualizing nitroreductase activity using magnetic resonance imaging.

Area of Science:

• Biomedical Engineering
• Molecular Imaging
• Synthetic Biology

Background:

• Bacterial nitroreductase is crucial for targeted cell ablation and gene-directed enzyme-prodrug therapy.
• Monitoring nitroreductase expression and activity is essential for these applications.
• Current optical imaging probes have limited penetration in deep tissues.

Purpose of the Study:

• To engineer a genetically encoded reporter for detecting nitroreductase activity using magnetic resonance imaging (MRI).
• To overcome the limitations of optical imaging for deep tissue applications.
• To enable noninvasive monitoring of nitroreductase-based genetic tools.

Main Methods:

• Molecular engineering of a reporter using human aquaporin fused to a dihydrofolate reductase (DHFR)-destabilizing domain.
• Utilizing nitroreductase-mediated cleavage of a caged prodrug to release trimethoprim, which stabilizes the aquaporin reporter.
• Optimizing signal induction through screening fusions and using an evolved DHFR variant.
• Validating the reporter's performance in diverse human cell types using diffusion-weighted MRI.

Main Results:

• A novel, genetically encoded MRI reporter system for bacterial nitroreductase activity was successfully developed.
• The reporter system demonstrated a direct, dose-dependent correlation between nitroreductase expression and MRI signal.
• The engineered sensor functions as a switchable, metal-free MRI reporter activated by nitroreductase activity.

Conclusions:

• Destabilized aquaporins serve as effective switchable, metal-free MRI reporters for monitoring nitroreductase activity.
• This technology facilitates noninvasive evaluation of nitroreductase-based genetic tools in intact deep tissues.
• The developed reporter has significant potential for advancing fundamental research and gene-directed enzyme-prodrug therapy.