Systematic Exploration of the Potential of HPQ as the Core Scaffold of Probes for in Vivo Imaging: A Case Study of FAP-Responsive Probes

The limited tissue penetration depth of optical probes based on 2-(2′-hydroxyphenyl)-4(3H)-quinazolinone (HPQ) hindered dynamic monitoring of the actual distribution and metabolism of them in deep tissues. Herein, using fibroblast activating protein (FAP)–activatable dual-modal probes as a prototype, we explored the potential of HPQ as a core scaffold for in vivo imaging probes via stepwise biological characterization. The tumor-to-muscle uptake ratios (T/M) of LWF-1, LWF-2, and LWF-3 were consistently above 4.05 during in vivo fluorescence imaging with sustained tumor visualization exceeding 48 h. All 68Ga-radiolabeled probes exhibited a maximum tumor uptake of up to 12.83 ± 9.37%ID/g in model mice, with the T/M of [68Ga]LWF-3 remaining above 4.89 throughout the PET scanning. The ex vivo biodistribution study revealed that [177Lu]LWF-3 reached the peak uptake in the tumor at 4 h postinjection (10.41 ± 0.68%ID/g). Accordingly, structural design and optimization governing their in vivo metabolism are critical for acquisition of ideal HPQ-based probes.