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Design, synthesis, and preclinical evaluation of a novel bifunctional macrocyclic chelator for theranostics of cancers

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Abstract

Purpose

This study was to design and synthesize a novel bifunctional chelator, named Dar, primarily validated by conjugating to tumor targeting motifs, labeled with radiometals, and performed preclinical evaluation of tumor imaging and cancer therapy in murine tumor models.

Method

The designed Dar was synthesized and characterized by X-ray crystallography, 1H/13C NMR, and mass spectrometry. Dar-PSMA-617 was conjugated and radiolabeled with 68Ga, 177Lu, and 89Zr. The in vivo behavior of 68 Ga/89Zr-labeled Dar-PSMA-617 were evaluated using micro-PET imaging and biodistribution from image quantitation and tissue radioactivity counting, with 68Ga/89Zr-labeled NOTA/DOTA/DFO-PSMA-617 analogs as controls, respectively. The [177Lu]-Dar-PSMA-617, with [177Lu]-DOTA-PSMA-617 as control, was evaluated in competitive cell uptake, tumor cell internalization, and efflux studies. The treatment efficacy of [177Lu]Lu-Dar-PSMA-617, with [177Lu]Lu-DOTA-PSMA-617 as control, was evaluated in PSMA-positive LNCaP tumor-bearing mice. In addition, the ability of Dar for radiolabeling nanobody was tested by conjugating Dar to KN035 nanobody. The resultant [89Zr]Zr-Dar-KN035 nanobody, with [89Zr]Zr-DFO-KN035 as control, was evaluated by micro-PET imaging and biodistribution in a mouse model bearing MC38&MC38-hPD-L1 colon cancer.

Results

68Ga, 89Zr, and 177Lu-radiolabeled Dar-PSMA-617 complexes were able to be produced under mild condition with high radiochemical yield and purity successfully. [177Lu]Lu-Dar-PSMA-617 had higher cellular uptake yet similar internalization and efflux properties in LNCaP cells, as compared to [177Lu]Lu-DOTA-PSMA-617. Micro-PET images demonstrated significantly higher tumor uptake of [68Ga]Ga-Dar-PSMA-617, than that of the analog [68Ga]Ga-DOTA-PSMA-617. The tumor uptake values of [68Ga]Ga-Dar-PSMA-617 at multiple time points are comparable to that of [68Ga]Ga-NOTA-PSMA-617, although a higher and persistently prolonged kidney retention was resulted in during the study period. The Dar chelator can also successfully mediate the radiolabeling with 89Zr, while the resultant [89Zr]Zr-Dar-PSMA-617 demonstrated a similar biodistribution with [89Zr]Zr-DFO-PSMA-617 measured at 96 h p.i. The treatment with [177Lu]Lu-Dar-PSMA-617 significantly inhibited the tumor growth, showing much better efficacy than that of [177Lu]Lu-DOTA-PSMA-617 at the same injected radioactivity and mass dose. Dar was covalently linked to KN035 nanobody and enabled radiolabeling with 89Zr in high yield and radiochemical purity at room temperature. The resultant [89Zr]Zr-Dar-KN035, with [89Zr]Zr-DFO-KN035 as control, demonstrated superior tumor uptake and detection capability in PET imaging studies.

Conclusion

The Dar, as a novel bifunctional chelator for medicating the labeling of radiometals onto tumor targeting carriers, was successfully synthesized and chemically characterized. Test radiolabeling, on PSMA-617 and a nanobody as tool targeting molecule carriers, demonstrated the Dar has potential as a universal bifunctional chelator for radiolabeling various radiometals (at least 68Ga, 177Lu, and 89Zr tested) commonly used for clinical imaging and therapy. Using a novel Dar chelator results in altered in vivo behavior of the carriers even though labeled with the same nuclide. This capability makes Dar an alternative to the existing choices for radiolabeling new carrier molecules with various radiometals, especially the radiometals with large radius.

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Acknowledgements

The authors wish to acknowledge the National Natural Science Foundation of China and China Postdoctoral Science Foundation for the support of this work.

Funding

This work was financially supported by the National Natural Science Foundation of China projects (Grant No. 11975123) and China Postdoctoral Science Foundation (Grant No. 2021T140321).

Author information

Authors and Affiliations

  1. Department of Nuclear Science and Technology, Nanjing University of Aeronautics and Astronautics, No. 29 Jiangjun Ave, Nanjing, 211016, People’s Republic of China

    Jianfeng Xu, Fei Cai, Changran Geng & Xiaobin Tang

  2. JYAMS PET Research and Development Limited, No. 568 Longmian Ave, Nanjing, 211100, People’s Republic of China

    Jianfeng Xu, Fei Cai, Zhigang Luo, Wenbin Fan, Juan Dai, Jingjing Cui, Shihong Li & Zheng Wang

  3. Department of Radiology and Imaging Sciences, Indiana University School of Medicine, 1345 West 16th Street, Room 112, Indianapolis, IN, 46202, USA

    Qihuang Zheng

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  1. Jianfeng Xu
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Contributions

XBT and ZW conceived the idea of the project. JFX wrote the manuscript in addition to designing, performing, and analyzing all experiments. JFX, FC, and WBF performed the experiments. JD, JJC, and SHL collected the information on animals. SHL and CRG assisted with data analysis. ZGL, CRG, QHZ, ZW, and XBT designed, supervised, and analyzed all experiments, in addition to assisting with manuscript preparation. All authors read and approved the final manuscript.

Corresponding authors

Correspondence to Zheng Wang or Xiaobin Tang.

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All animal studies were performed in accordance with the protocols provided in the Guide for the Care and Use of Medical Laboratory Animals (Ministry of Health, China).

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The authors declare no competing interests.

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Xu, J., Cai, F., Luo, Z. et al. Design, synthesis, and preclinical evaluation of a novel bifunctional macrocyclic chelator for theranostics of cancers. Eur J Nucl Med Mol Imaging 49, 2618–2633 (2022). https://doi.org/10.1007/s00259-022-05750-8

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  • DOI: https://doi.org/10.1007/s00259-022-05750-8

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