Tolmachev Vladimir, Papalanis Eleftherios, Bezverkhniaia Ekaterina A, Rosly Alia Hani, Vorobyeva Anzhelika, Orlova Anna, Carlqvist Matilda, Frejd Fredrik Y, Oroujeni Maryam
Department of Immunology, Genetics and Pathology, Uppsala University, 751 85 Uppsala, Sweden.
Department of Medicinal Chemistry, Uppsala University, 751 83 Uppsala, Sweden.
ACS Pharmacol Transl Sci. 2025 Feb 19;8(3):706-717. doi: 10.1021/acsptsci.4c00539. eCollection 2025 Mar 14.
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 Tc-labeled probes, AC12 and an affinity-matured SYNT179 using a [Tc]Tc-GGGC label. This study aimed to evaluate whether the use of a residualizing In-based label provides better imaging contrast compared with a nonresidualizing label. To do that, SYNT179 and AC12-GGGC Affibody molecules were labeled with In 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 In-labeled conjugates to B7-H3-expressing living cells were higher compared with the affinities of the Tc-labeled variants. In mice with B7-H3-expressing xenografts, the tumor uptake of In-labeled proteins (3.6 ± 0.3 and 1.8 ± 0.5%ID/g for [In]In-SYNT179-DOTA and [In]In-AC12-DOTA, respectively) was significantly ( < 0.05, ANOVA) higher than those for Tc-labeled counterparts (1.6 ± 0.2%ID/g and 0.8 ± 0.2%ID/g for [Tc]Tc-SYNT179 and [Tc]Tc-AC12-GGGC, respectively). The best variant, [In]In-SYNT179-DOTA, provided a tumor-to-blood ratio of 31.1 ± 2.9, which was twice higher than that for [Tc]Tc-SYNT179 and 7-fold higher than that for [Tc]Tc-AC12-GGGC. Both In-labeled Affibody molecules had higher renal retention compared with Tc-labeled ones, but the hepatobiliary excretion of In-labeled proteins was appreciably lower, potentially improving the imaging of abdominal metastases. Overall, [In]In-SYNT179-DOTA is the most promising tracer for visualization of B7-H3 expression.
免疫检查点蛋白B7-H3（CD276）在多种癌症中过表达，是恶性肿瘤治疗的一个有吸引力的靶点。使用工程化支架蛋白（如亲合素分子）对B7-H3表达进行放射性核素分子成像，是选择B7-H3靶向治疗潜在应答者的一种有前景的策略。使用两种锝标记的探针AC12和使用[Tc]Tc-GGGC标记的亲和力成熟的SYNT179，证明了B7-H3成像的可行性。本研究旨在评估与非残留标记相比，使用残留性铟基标记是否能提供更好的成像对比度。为此，使用（4,10-双羧甲基-7-{[2-(2,5-二氧代-3-硫代吡咯烷-1-基)-乙基氨基甲酰基]-甲基}-1,4,7,10-四氮杂环十二烷基）-乙酸（马来酰亚胺-DOTA）螯合剂将SYNT179和AC12-GGGC亲合素分子用铟标记，该螯合剂位点特异性地偶联到亲合素分子的C末端。与锝标记的变体相比，铟标记的结合物与表达B7-H3的活细胞的结合亲和力更高。在表达B7-H3的异种移植小鼠中，铟标记蛋白的肿瘤摄取（[In]In-SYNT179-DOTA和[In]In-AC12-DOTA分别为3.6±0.3和1.8±0.5%ID/g）显著（<0.05，方差分析）高于锝标记对应物（[Tc]Tc-SYNT179和[Tc]Tc-AC12-GGGC分别为1.6±0.2%ID/g和0.8±0.2%ID/g）。最佳变体[In]In-SYNT179-DOTA的肿瘤与血液比值为31.1±2.9，是[Tc]Tc-SYNT179的两倍，是[Tc]Tc-AC12-GGGC的7倍。与锝标记的亲合素分子相比，两种铟标记的亲合素分子的肾脏滞留率更高，但铟标记蛋白的肝胆排泄明显更低，这可能会改善腹部转移灶的成像。总体而言，[In]In-SYNT179-DOTA是可视化B7-H3表达最有前景的示踪剂。
