AND-gate contrast agents for enhanced fluorescence-guided surgery

The surgical resection of tumours requires the precise location and definition of the margins between lesions and normal tissue. However, this is made difficult by irregular margin borders. Although molecularly targeted optical contrast agents can be used to define tumour margins during surgery in real time, the selectivity of the contrast agents is often limited by the target being expressed in both healthy and tumour tissues. Here, we show that AND-gate optical imaging probes requiring the processing of two substrates by multiple tumour-specific enzymes produce a fluorescent signal with significantly improved specificity and sensitivity to tumour tissue. We evaluated the performance of the probes in mouse models of mammary tumours and of metastatic lung cancer, and during fluorescence-guided robotic surgery. Imaging probes relying on multivariate activation to selectively target complex patterns of enzymatic activity should be useful in disease detection, treatment and monitoring.