The 4T1 murine breast cancer model was generated as previously described [13 (link), 29 (link)]. PET and PET/CT scans of tumor-bearing female BALB/c mice (6–8 weeks old, tumor diameter 5–8 mm) were performed using an Inveon microPET/microCT rodent model scanner (Siemens Medical Solutions USA, Inc.). Each mouse was intravenously injected with 5–10 MBq of 61/64Cu-NOTA-TRC105-Fab and 5 or 10 minute static PET scans were performed at various time points post-injection (p.i.). The images were reconstructed using a maximum a posteriori (MAP) algorithm, with no attenuation or scatter correction. Region-of-interest (ROI) analysis of each PET scan was performed using vendor software (Inveon Research Workplace [IRW]) on decay-corrected whole-body images as described previously [30 (link), 31 (link)], to calculate the percentage injected dose per gram of tissue (%ID/g) values for the 4T1 tumor and several major organs.
Blocking studies were carried out to evaluate CD105 specificity of 64Cu-NOTA-TRC105-Fab in vivo, where a group of 4 mice was each injected with 2 mg of TRC105 within 1 h before 64Cu-NOTA-TRC105-Fab administration. After the last PET scans at 24 h p.i., mice were euthanized and the blood, 4T1 tumor, and major organs/tissues were collected and wet-weighed. The radioactivity in the tissue was measured using a Cobra II gamma-counter (Perkin-Elmer) and presented as %ID/g. The 4T1 tumor, liver, kidney (i.e. tissues with significant uptake of 61/64Cu-NOTA-TRC105-Fab), and muscle were also frozen and sectioned for histological analysis.