Evaluation of the first ⁴⁴Sc-labeled Affibody molecule for imaging of HER2-expressing tumors

Introduction Affibody molecules are small (58 amino acids) high-affinity proteins based on a tri-helix non-immunoglobulin scaffold. A clinical study has demonstrated that PET imaging using Affibody molecules labeled with ⁶⁸Ga (T_½ = 68 min) can visualize metastases of breast cancer expressing human epidermal growth factor receptor type 2 (HER2) and provide discrimination between tumors with high and low expression level. This may help to identify breast cancer patients benefiting from HER2-targeting therapies. The best discrimination was at 4 h post injection. Due to longer half-life, a positron-emitting radionuclide ⁴⁴Sc (T_½ = 4.04 h) might be a preferable label for Affibody molecules for imaging at several hours after injection. Methods A synthetic second-generation anti-HER2 Affibody molecule Z_HER2:2891 was labeled with ⁴⁴Sc via a DOTA-chelator conjugated to the N-terminal amino group. Binding specificity, affinity and cellular processing ⁴⁴Sc-DOTA-Z_HER2:2891 and ⁶⁸Ga-DOTA-Z_HER2:2891 were compared in vitro using HER2-expressing cells. Biodistribution and imaging properties of ⁴⁴Sc-DOTA-Z_HER2:2891 and ⁶⁸Ga-DOTA-Z_HER2:2891 were evaluated in Balb/c nude mice bearing HER2-expression xenografts. Results The labeling yield of 98 ± 2% and specific activity of 7.8 GBq/μmol were obtained. The conjugate demonstrated specific binding to HER2-expressing SKOV3.ip cells in vitro and to SKOV3.ip xenografts in nude mice. The distribution of radioactivity at 3 h post injection was similar for ⁴⁴Sc-DOTA-Z_HER2:2891 and ⁶⁸Ga-DOTA-Z_HER2:2891, but the blood clearance of the ⁴⁴Sc-labeled variant was slower and the tumor-to-blood ratio was reduced (15 ± 2 for ⁴⁴Sc-DOTA-Z_HER2:2891 vs 46 ± 9 for ⁶⁸Ga-DOTA-Z_HER2:2891). At 6 h after injection of ⁴⁴Sc-DOTA-Z_HER2:2891 the tumor uptake was 8 ± 2% IA/g and the tumor-to-blood ratio was 51 ± 8. Imaging using small-animal PET/CT demonstrated that ⁴⁴Sc-DOTA-Z_HER2:2891 provides specific and high-contrast imaging of HER2-expressing xenografts. Conclusion The ⁴⁴Sc- DOTA-Z_HER2:2891 Affibody molecule is a promising probe for imaging of HER2-expression in malignant tumors.