All animal studies were conducted under a protocol approved by the University of Wisconsin Institutional Animal Care and Use Committee. PET and PET/CT scans at various time points p.i. using a microPET/microCT Inveon rodent model scanner (Siemens Medical Solutions USA, Inc.), image reconstruction, and ROI analysis of the PET data were performed similar as described previously (see Supporting Information for details).49 (link), 50 (link) Quantitative PET data were presented as percentage injected dose per gram of tissue (%ID/g).
4T1 murine breast tumor-bearing mice, a fast-growing tumor model with high CD105 expression on the tumor vasculature,41 (link), 42 (link), 50 (link) were each injected with 5–10 MBq of 64Cu-NOTA-GO-TRC105 or 64Cu-NOTA-GO via tail vein before serial PET scans. Another group of four 4T1 tumor-bearing mice were each injected with 2 mg of unlabeled TRC105 at 2 h before 64Cu-NOTA-GO-TRC105 administration to evaluate the CD105 specificity of 64Cu-NOTA-GO-TRC105 in vivo (i.e. blocking experiment). To generate the 4T1 tumor model, 4- to 5-week-old female Balb/c mice were purchased from Harlan (Indianapolis, IN, USA) and tumors were established by subcutaneously injecting 2×106 cells, suspended in 100 μl of 1:1 mixture of RPMI 1640 and Matrigel (BD Biosciences, Franklin Lakes, NJ, USA), into the front flank of mice. The tumor sizes were monitored every other day and the animals were subjected to in vivo experiments when the tumor diameter reached 5–8 mm.
After the last PET scans at 48 h p.i., biodistribution studies were carried out to confirm that the %ID/g values based on PET imaging truly represented the radioactivity distribution in tumor-bearing mice. In addition, separate groups of four 4T1 tumor-bearing mice were each intravenously injected with 64Cu-NOTA-GO-TRC105 or 64Cu-NOTA-GO and euthanized at 3 h p.i. (when tumor uptake was at the peak based on PET results) for biodistribution studies. Mice were euthanized and blood, 4T1 tumor, and major organs/tissues were collected and wet-weighed. The radioactivity in the tissue was measured using a gamma-counter (Perkin Elmer) and presented as %ID/g (mean ± SD).
4T1 murine breast tumor-bearing mice, a fast-growing tumor model with high CD105 expression on the tumor vasculature,41 (link), 42 (link), 50 (link) were each injected with 5–10 MBq of 64Cu-NOTA-GO-TRC105 or 64Cu-NOTA-GO via tail vein before serial PET scans. Another group of four 4T1 tumor-bearing mice were each injected with 2 mg of unlabeled TRC105 at 2 h before 64Cu-NOTA-GO-TRC105 administration to evaluate the CD105 specificity of 64Cu-NOTA-GO-TRC105 in vivo (i.e. blocking experiment). To generate the 4T1 tumor model, 4- to 5-week-old female Balb/c mice were purchased from Harlan (Indianapolis, IN, USA) and tumors were established by subcutaneously injecting 2×106 cells, suspended in 100 μl of 1:1 mixture of RPMI 1640 and Matrigel (BD Biosciences, Franklin Lakes, NJ, USA), into the front flank of mice. The tumor sizes were monitored every other day and the animals were subjected to in vivo experiments when the tumor diameter reached 5–8 mm.
After the last PET scans at 48 h p.i., biodistribution studies were carried out to confirm that the %ID/g values based on PET imaging truly represented the radioactivity distribution in tumor-bearing mice. In addition, separate groups of four 4T1 tumor-bearing mice were each intravenously injected with 64Cu-NOTA-GO-TRC105 or 64Cu-NOTA-GO and euthanized at 3 h p.i. (when tumor uptake was at the peak based on PET results) for biodistribution studies. Mice were euthanized and blood, 4T1 tumor, and major organs/tissues were collected and wet-weighed. The radioactivity in the tissue was measured using a gamma-counter (Perkin Elmer) and presented as %ID/g (mean ± SD).
