PET imaging and biodistribution studies

PET scans were performed using a preclinical Inveon microPET/CT (Siemens Medical Solutions USA, Inc.). Each tumor-bearing mouse was injected with 5–10 MBq of ⁸⁹Zr-Df-ATN-291 into its tail vein and subjected to static PET scans (40 million events per scan) at various time points post-injection (p.i.). The images were reconstructed using a three-dimensional ordered subset expectation maximization (3D-OSEM) algorithm, with no attenuation or scatter correction. For each microPET scan, Inveon Research Workshop (IRW, v4.2.0.8) was used to superimpose three-dimensional (3D) regions-of-interest (ROIs) on the tumor and major organs in the decay-corrected whole-body images. By adopting a tissue density of 1 g/mL, the radioactivity in each ROI volume was converted to MBq/g using a conversion factor and then divided by the total administered radioactivity to obtain a percentage of injected dose per gram of tissue (%ID/g) for each organ/tissue.

Biodistribution studies were carried out to confirm that the quantitative tracer uptake values based on PET imaging truly represented the radioactivity distribution in tumor-bearing mice. After the last PET scans at 120 h p.i., mice were euthanized and blood, tumors, and major organs/tissues were collected and wet-weighed. The radioactivity in each collected sample was measured using a WIZARD² automatic gamma-counter (Perkin-Elmer) and recorded as %ID/g (mean ± SD). The tumors were also frozen for histological analysis.