Imaging of ROR1 expression in tumors using radiolabeled affibody molecules
- 1Department of Protein Science, School of Engineering Sciences in Chemistry, Biotechnology and Health, KTH Royal Institute of Technology, 100 44 Stockholm, Sweden
- 2Department of Immunology, Genetics and Pathology, Uppsala University, 751 85 Uppsala, Sweden
- 3Department of Medicinal Chemistry, Uppsala University, 751 23 Uppsala, Sweden
Abstract
Purpose: Receptor tyrosine kinase-like orphan receptor 1 (ROR1) is an attractive molecular target for anti-tumor therapies undergoing late-stage clinical development. Quantitative imaging of ROR1 expression could enable identification of patients likely to respond to such treatments. This study aimed to assess the feasibility of specific in vivo imaging of ROR1 using radiolabeled affibody molecules.
Methods: The affibody molecule ZROR1:A10, with high affinity for human (4 nM) and murine (2 nM) ROR1, was labeled with indium-111 using a DOTA chelator. In vitro binding specificity, affinity, and cellular processing were evaluated using a panel of ROR1-expressing cell lines. The affibody [111In]In-DOTA-Zcov19s, which does not bind ROR1, served as a nonspecific control. The biodistribution of [111In]In-DOTA-ZROR1:A10 and [111In]In-DOTA-Zcov19s was measured in immunodeficient mice bearing ROR1-positive tumors, with ROR1-negative Ramos xenografts used as an additional specificity control.
Results: [111In]In-DOTA-ZROR1:A10 bound specifically to ROR1-expressing cells, with significantly higher uptake than [111In]In-DOTA-Zcov19s. The apparent equilibrium dissociation constant for ROR1 binding in vitro was 1–4 nM. Tumor uptake of [111In]In-DOTA-ZROR1:A10 in MDA-MB-468 xenografts was saturable and significantly (p < 0.05) higher than in Ramos tumors, as well as higher than uptake of the control affibody. At 4 h post‑injection of 1 µg (50 µg/kg), tumor‑to‑blood, tumor‑to‑bone, and tumor‑to‑muscle ratios were 14 ± 6, 19 ± 6, and 11 ± 1, respectively. Micro-SPECT/CT imaging clearly visualized ROR1‑positive tumors and discriminated them from ROR1‑negative xenografts.
Conclusion: The preclinical evaluation shows that visualization of ROR1-expressing tumors using a radiolabeled affibody molecule is feasible.
Supplementary Information: The online version contains supplementary material available at https://doi.org/10.1007/s00259-026-08106-8.
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BibTeX
@article{westerberg2026imaging,
title = {{Imaging of ROR1 expression in tumors using radiolabeled affibody molecules}},
author = {Cornelia Westerberg and Tianqi Xu and Maria Marth and Moeen Ud-Din and Russol Abushaia and Tilde Hagelin and Stefan Ståhl and Anna Orlova and Vladimir Tolmachev and John Löfblom and Anzhelika Vorobyeva},
journal = {European Journal of Nuclear Medicine and Molecular Imaging},
year = {2026},
month = aug,
doi = {10.1007/s00259-026-08106-8},
url = {https://doi.org/10.1007/s00259-026-08106-8},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Imaging of ROR1 expression in tumors using radiolabeled affibody molecules
AU - Cornelia Westerberg
AU - Tianqi Xu
AU - Maria Marth
AU - Moeen Ud-Din
AU - Russol Abushaia
AU - Tilde Hagelin
AU - Stefan Ståhl
AU - Anna Orlova
AU - Vladimir Tolmachev
AU - John Löfblom
AU - Anzhelika Vorobyeva
PY - 2026
DA - 2026/08/01
JO - European Journal of Nuclear Medicine and Molecular Imaging
DO - 10.1007/s00259-026-08106-8
UR - https://doi.org/10.1007/s00259-026-08106-8
AB - Purpose: Receptor tyrosine kinase-like orphan receptor 1 (ROR1) is an attractive molecular target for anti-tumor therapies undergoing late-stage clinical development. Quantitative imaging of ROR1 expression could enable identification of patients likely to respond to such treatments. This study aimed to assess the feasibility of specific in vivo imaging of ROR1 using radiolabeled affibody molecules. Methods: The affibody molecule ZROR1:A10, with high affinity for human (4 nM) and murine (2 nM) ROR1, was labeled with indium-111 using a DOTA chelator. In vitro binding specificity, affinity, and cellular processing were evaluated using a panel of ROR1-expressing cell lines. The affibody [111In]In-DOTA-Zcov19s, which does not bind ROR1, served as a nonspecific control. The biodistribution of [111In]In-DOTA-ZROR1:A10 and [111In]In-DOTA-Zcov19s was measured in immunodeficient mice bearing ROR1-positive tumors, with ROR1-negative Ramos xenografts used as an additional specificity control. Results: [111In]In-DOTA-ZROR1:A10 bound specifically to ROR1-expressing cells, with significantly higher uptake than [111In]In-DOTA-Zcov19s. The apparent equilibrium dissociation constant for ROR1 binding in vitro was 1–4 nM. Tumor uptake of [111In]In-DOTA-ZROR1:A10 in MDA-MB-468 xenografts was saturable and significantly (p < 0.05) higher than in Ramos tumors, as well as higher than uptake of the control affibody. At 4 h post‑injection of 1 µg (50 µg/kg), tumor‑to‑blood, tumor‑to‑bone, and tumor‑to‑muscle ratios were 14 ± 6, 19 ± 6, and 11 ± 1, respectively. Micro-SPECT/CT imaging clearly visualized ROR1‑positive tumors and discriminated them from ROR1‑negative xenografts. Conclusion: The preclinical evaluation shows that visualization of ROR1-expressing tumors using a radiolabeled affibody molecule is feasible. Supplementary Information: The online version contains supplementary material available at https://doi.org/10.1007/s00259-026-08106-8.
SN - 1619-7070
SN - 1619-7089
ER -