⁶⁴Cu-DOTA-Dara detects MM cell dissemination in vivo. (A) The radiolabeled antibody ⁶⁴Cu-DOTA-Dara (3.7 MBq, 10 µg) was administered to 3 mice that had been transplanted by IV injection with a human MM cell line (MM.1S GFP⁺/Luc⁺) before (7 days post-IV) and after (21 days post-IV) signs of MM bone metastasis as detected by BLI. The mice were also imaged with ⁶⁴Cu-DOTA-Dara radiolabeled antibody on the following days, and PET imaging identified MM cell dissemination in the bone (yellow boxes), which overlap with the BLI signals in the same mouse. (B) ⁶⁴Cu-DOTA-Dara (3.7 MBq, 10 µg) was also administered to 2 mice that had been transplanted by IV injection with U266 Luc⁺ MM cells, when clear signs (5 weeks post-IV) of MM bone engraftment were detected by BLI. The mice were imaged with ⁶⁴Cu-DOTA-Dara radiolabeled antibody after 1 day from the radiolabeled Dara injection (see also supplemental Figure 4B). Of note, no sign of engraftment was observed by PET/CT scan. (C) ⁶⁴Cu-DOTA-trastuzumab (⁶⁴Cu-DOTA-trast) (3.7 MBq, 10 µg) was administrated to 3 MM.1S GFP⁺/Luc⁺–engrafted mice, as assessed by BLI imaging, and a PET/CT scan was performed 1 day after ⁶⁴Cu-DOTA-trast injection. No bone signals were observed, but only high background associated with the circulating radiolabeled antibody. (D-E) Mice bearing CD38⁺ MM cells (MM.1S GFP⁺/Luc⁺) with a high level of engraftment (see also supplemental Figure 4C), as assessed by BLI images, that were injected with ⁶⁴Cu-DOTA-Dara (3.7 MBq, 10 µg) without (D) and with (E) unlabeled Dara (500 µg) and imaged at day 1 by PET/CT. Almost no signs of engraftments were detected in the presence of cold Dara, independent of the BLI signals. (F) ⁶⁴Cu-DOTA-Dara PET/CT-positive mice (MM.1S GFP⁺/Luc⁺–engrafted mice: 4) and negative mice (tumor free mice: 4); ⁶⁴Cu-DOTA-trast–treated mice (3); unlabeled Dara (Block) treated mice (2); and U266-engrafted mice (2) were sacrificed, and organs were harvested at 24 hours and used for γ counting to determine their radioactive content, as is presented in the bar graphs as a percentage of the injected dose per gram (%ID/g). See also supplemental Figure 5A.