Comparative Preclinical Evaluation of the Tumor-Targeting Properties of Radioiodine and Technetium-Labeled Designed Ankyrin Repeat Proteins for Imaging of Epidermal Growth Factor Receptor Expression in Malignant Tumors
Radionuclide molecular imaging of epidermal growth factor receptor (EGFR) expression might permit the selection of patients for EGFR-targeting therapies. Designed ankyrin repeat protein (DARPin) E01 with a high affinity to the ectodomain III of the EGFR is a possible EGFR imaging probe. The goal of this study was to evaluate the potential of radiolabeled DARPin E01 for in vivo imaging of EGFR. DARPin E01 containing the (HE)<sub>3</sub>-tag was site-specifically labeled with a residualizing <sup>99m</sup>Tc (using <sup>99m</sup>Tc]Tc(CO)<sub>3</sub>). Two methods providing non-residualizing <sup>123</sup>I labels, direct electrophilic radioiodination and indirect radioiodination using [<sup>123</sup>I]I-<i>para</i>-iodobenzoate (PIB), were tested. [<sup>99m</sup>Tc]Tc-(HE)<sub>3</sub>-E01 and [<sup>123</sup>I]I-(HE)<sub>3</sub>-E01-PIB preserved specific binding to EGFR-expressing cells and affinity in the single-digit nanomolar range. Direct labeling with <sup>123</sup>I resulted in a substantial loss of binding. In vitro cellular processing studies showed that both [<sup>99m</sup>Tc]Tc-(HE)<sub>3</sub>-E01 and [<sup>123</sup>I]I-(HE)<sub>3</sub>-E01-PIB had rapid binding and relatively slow internalization. Evaluation of [<sup>99m</sup>Tc]Tc-(HE)<sub>3</sub>-E01 biodistribution in normal CD1 mice showed that its hepatic uptake was non-saturable, suggesting that this tracer does not bind to murine EGFR. A side-by-side comparison of biodistribution and tumor targeting of [<sup>99m</sup>Tc]Tc-(HE)<sub>3</sub>-E01 and [<sup>123</sup>I]I-(HE)<sub>3</sub>-E01-PIB was performed in Nu/j mice bearing EGFR-positive A-431 and EGFR-negative Ramos human cancer xenografts. Both radiolabeled DARPins demonstrated EGFR-specific tumor uptake. However, [<sup>123</sup>I]I-(HE)<sub>3</sub>-E01-PIB had appreciably lower uptake in normal organs compared to [<sup>99m</sup>Tc]Tc-(HE)<sub>3</sub>-E01, which provided significantly (<i>p</i> < 0.05) higher tumor-to-organ ratios. Gamma-camera imaging confirmed that [<sup>123</sup>I]I-(HE)<sub>3</sub>-E01-PIB demonstrated a higher imaging contrast in preclinical models than [<sup>99m</sup>Tc]Tc-(HE)<sub>3</sub>-E01. In conclusion, DARPin (HE)<sub>3</sub>-E01 labeled using a non-residualizing [<sup>123</sup>I]I-<i>para</i>-iodobenzoate (PIB) label is the preferred radiotracer for in vivo imaging of EGFR expression in cancer.
Authors
- Rustam Ziganshin (ORCID: https://orcid.org/0000-0002-7931-519X)
- Evgenii Plotnikov (ORCID: https://orcid.org/0000-0002-4374-6422)
- Alexey Schulga (ORCID: https://orcid.org/0000-0003-3425-0828)
- M. V. Tretyakova (ORCID: https://orcid.org/0000-0002-3628-0804)
- Lutfi Aditya Hasnowo (ORCID: https://orcid.org/0000-0001-5614-9603)
- М. В. Белоусов (ORCID: https://orcid.org/0000-0002-2153-7945)
- Sergey M. Deyev (ORCID: https://orcid.org/0000-0002-3952-0631)
- Е. В. Коновалова (ORCID: https://orcid.org/0000-0002-4187-376X)
- Feruza Yuldasheva (ORCID: https://orcid.org/0000-0001-9907-9334)
- Vladimir Tolmachev (ORCID: https://orcid.org/0000-0002-6122-1734)
- Ruslan Varvashenya (ORCID: https://orcid.org/0009-0002-4282-3198)
- M. S. Larkina (ORCID: https://orcid.org/0000-0003-1176-2441)
- Roman Zelchan
- Gleb Yanovich
Institutions
- Uppsala University (SE)
- Russian Academy of Sciences (RU)
- National Research Tomsk State University (RU)
- Siberian State Medical University (RU)
- Tomsk Polytechnic University (RU)
- Institute of Bioorganic Chemistry (RU)
- Tomsk National Research Medical Center (RU)
- National Nuclear Energy Agency of Indonesia (ID)
Publication Details
- Journal
- International Journal of Molecular Sciences
- Published
- 2025-10-31
- DOI
- https://doi.org/10.3390/ijms262110609
- Citations
- 1
- Primary Topic
- Radiopharmaceutical Chemistry and Applications
- Type
- article
- Field-Weighted Citation Impact
- 1.36