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Impact of Radiometal Chelates on In Vivo Visualization of Immune Checkpoint Protein Using Radiolabeled Affibody
Vladimir Tolmachev1, Eleftherios Papalanis1, Ekaterina A Bezverkhniaia2
1Department of Immunology, Genetics and Pathology, Uppsala University, 751 85 Uppsala, Sweden.
Abstract:
The immune checkpoint protein B7-H3 (CD276) is overexpressed in various cancers and is an attractive target for the treatment of malignant tumors. Radionuclide molecular imaging of B7-H3 expression using engineered scaffold proteins such as Affibody molecules is a promising strategy for the selection of potential responders to B7-H3-targeted therapy. Feasibility of B7-H3 imaging was demonstrated using two 99mTc-labeled probes, AC12 and an affinity-matured SYNT179 using a [99mTc]Tc-GGGC label. This study aimed to evaluate whether the use of a residualizing 111In-based label provides better imaging contrast compared with a nonresidualizing label. To do that, SYNT179 and AC12-GGGC Affibody molecules were labeled with 111In using (4,10-bis-carboxymethyl-7-{[2-(2,5-dioxo-3-thioxo-pyrrolidin-1-yl)-ethylcarbamoyl]-methyl}-1,4,7,10-tetraaza-cyclododec-1-yl)-acetic acid (maleimide-DOTA) chelator, site-specifically coupled to the C-terminus of Affibody molecules. The binding affinities of the 111In-labeled conjugates to B7-H3-expressing living cells were higher compared with the affinities of the 99mTc-labeled variants. In mice with B7-H3-expressing xenografts, the tumor uptake of 111In-labeled proteins (3.6 ± 0.3 and 1.8 ± 0.5%ID/g for [111In]In-SYNT179-DOTA and [111In]In-AC12-DOTA, respectively) was significantly (p < 0.05, ANOVA) higher than those for 99mTc-labeled counterparts (1.6 ± 0.2%ID/g and 0.8 ± 0.2%ID/g for [99mTc]Tc-SYNT179 and [99mTc]Tc-AC12-GGGC, respectively). The best variant, [111In]In-SYNT179-DOTA, provided a tumor-to-blood ratio of 31.1 ± 2.9, which was twice higher than that for [99mTc]Tc-SYNT179 and 7-fold higher than that for [99mTc]Tc-AC12-GGGC. Both 111In-labeled Affibody molecules had higher renal retention compared with 99mTc-labeled ones, but the hepatobiliary excretion of 111In-labeled proteins was appreciably lower, potentially improving the imaging of abdominal metastases. Overall, [111In]In-SYNT179-DOTA is the most promising tracer for visualization of B7-H3 expression.
Insights
This study demonstrates that Indium-111 (¹¹¹In)-labeled Affibody molecules offer superior imaging of B7-H3 (CD276) expression in cancer compared to Technetium-99m (⁹⁹mTc)-labeled versions. The ¹¹¹In-SYNT179-DOTA tracer showed significantly higher tumor uptake and contrast, making it promising for targeted cancer therapy selection.
Area of Science:
- Nuclear Medicine
- Oncology
- Radiopharmaceutical Chemistry
Background:
- B7-H3 (CD276) is overexpressed in various cancers, making it a significant therapeutic target.
- Radionuclide molecular imaging using engineered scaffold proteins like Affibody molecules is crucial for identifying patients who may benefit from B7-H3-targeted therapies.
- Previous studies demonstrated feasibility of B7-H3 imaging with ⁹⁹mTc-labeled probes.
Purpose of the Study:
- To evaluate if a residualizing ¹¹¹In-based label enhances imaging contrast for B7-H3 expression compared to a nonresidualizing label.
- To compare the performance of ¹¹¹In-labeled Affibody molecules (SYNT179 and AC12-GGGC) with their ⁹⁹mTc-labeled counterparts.
Main Methods:
- SYNT179 and AC12-GGGC Affibody molecules were site-specifically labeled with ¹¹¹In using a maleimide-DOTA chelator.
- Binding affinities of ¹¹¹In- and ⁹⁹mTc-labeled conjugates were assessed using B7-H3-expressing cells.
- In vivo imaging studies were conducted in mice bearing B7-H3-expressing xenografts to measure tumor uptake and biodistribution.
Main Results:
- The binding affinities of ¹¹¹In-labeled Affibody molecules were higher than their ⁹⁹mTc-labeled variants.
- Tumor uptake of ¹¹¹In-labeled proteins ([¹¹¹In]In-SYNT179-DOTA and [¹¹¹In]In-AC12-DOTA) was significantly higher (p < 0.05) than ⁹⁹mTc-labeled counterparts.
- [¹¹¹In]In-SYNT179-DOTA achieved a tumor-to-blood ratio of 31.1 ± 2.9, which was double that of [⁹⁹mTc]Tc-SYNT179 and sevenfold higher than [⁹⁹mTc]Tc-AC12-GGGC. Hepatobiliary excretion was lower for ¹¹¹In-labeled probes.
Conclusions:
- The residualizing ¹¹¹In label significantly improves imaging contrast for B7-H3 expression compared to the nonresidualizing ⁹⁹mTc label.
- [¹¹¹In]In-SYNT179-DOTA demonstrates superior performance, showing high tumor uptake and favorable biodistribution.
- [¹¹¹In]In-SYNT179-DOTA is identified as the most promising radiotracer for visualizing B7-H3 expression in the context of targeted cancer therapy.
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