Simple SummaryThe receptor HER2 is overexpressed in some breast cancers. Tumours with a high HER2 expression can be successfully treated with the antibodies trastuzumab and pertuzumab. The radionuclide imaging of HER2 in disseminated cancer could help to select patients for treatment using these antibodies. Novel radiolabelled small-sized tracers, scaffold proteins, have shown excellent imaging properties in preclinical studies. The scaffold proteins [99mTc]Tc-ADAPT6 and DARPin [99mTc]Tc-(HE)3-G3 have been found to be safe in Phase I clinical trials. They showed promising results in the imaging of HER2. In this study, we compared the distribution of both tracers in the same patients with breast cancer to evaluate whether one of them has any decisive advantage. We found that both tracers provide an excellent visualization of tumours, but the accumulation of [99mTc]Tc-ADAPT6 in tumours is higher. The data from this study are essential for researchers developing imaging agents.AbstractPrevious Phase I clinical evaluations of the radiolabelled scaffold proteins [99mTc]Tc-ADAPT6 and DARPin [99mTc]Tc-(HE)3-G3 in breast cancer patients have demonstrated their safety and indicated their capability to discriminate between HER2-positive and HER2-negative tumours. The objective of this study was to compare the imaging of HER2-positive tumours in the same patients using [99mTc]Tc-ADAPT6 and [99mTc]Tc-(HE)3-G3. Eleven treatment-naïve female patients (26–65 years) with HER2-positive primary and metastatic breast cancer were included in the study. Each patient was intravenously injected with [99mTc]Tc-ADAPT6, followed by an [99mTc]Tc-(HE)3-G3 injection 3–4 days later and chest SPECT/CT was performed. All primary tumours were clearly visualized using both tracers. The uptake of [99mTc]Tc-ADAPT6 in primary tumours (SUVmax = 4.7 ± 2.1) was significantly higher (p< 0.005) than the uptake of [99mTc]Tc-(HE)3-G3 (SUVmax = 3.5 ± 1.7). There was no significant difference in primary tumour-to-contralateral site values for [99mTc]Tc-ADAPT6 (15.2 ± 7.4) and [99mTc]Tc-(HE)3-G3 (19.6 ± 12.4). All known lymph node metastases were visualized using both tracers. The uptake of [99mTc]Tc-ADAPT6 in all extrahepatic soft tissue lesions was significantly (p< 0.0004) higher than the uptake of [99mTc]Tc-(HE)3-G3. In conclusion, [99mTc]Tc-ADAPT6 and [99mTc]Tc-(HE)3-G3 are suitable for the visualization of HER2-positive breast cancer. At the selected time points, [99mTc]Tc-ADAPT6 has a significantly higher uptake in soft tissue lesions, which might be an advantage for the visualization of small metastases.