Poster Presentation 21st International Conference on Biological Inorganic Chemistry 2025

Light-activated ortho-nitrobenzyl alcohols for the conjugation of 89Zirconium-DFO to onartuzumab for tumor imaging (#509)

James W Southwell 1 , Cesare Berton 1 , Simon Klingler 1 , Jason P Holland 1
  1. University of Zurich, Zurich, Switzerland

89Zr-radiolabelled monoclonal antibodies (mAbs) play a central role in the diagnosis and monitoring of cancer treatment via non-invasive positron emission tomography (PET) imaging. Methods that provide access to 89Zr-mAbs in a simple, fast, and efficient manner from the native mAb are highly desirable. To this end, we report the synthesis of a desferrioxamine (DFO) ligand decorated with an ortho-nitrobenzyl alcohol (NBA)1 motif (DFO-NBA) for light-activated conjugation to the gastric cancer-targeting mAb onartuzumab2 for in vitro and in vivo characterization, Figure 1.

DFO-NBA was synthesized in three steps. 89Zr-radiolabeling was followed by TLC and HPLC. Photo-conjugation to the mAb was carried out using 365 nm light for 15 min under aqueous conditions (pH 8.1, 23 oC), and followed by TLC and size-exclusion chromatography (SEC). Tumor specificity was determined using cellular binding assays in vitro. The radiotracer pharmacokinetic profile and tumor targeting was then assessed by PET imaging in female athymic nude mice bearing subcutaneous MKN-45 xenografts. Biodistribution experiments were performed after 72 h. The tumor specificity of 89ZrDFO-NBA-onartuzumab was assessed in vivo by competitive inhibition (blocking) studies.

89Zr-DFO-NBA was synthesized with a radiochemical conversion (RCC) >99%. Photo-conjugation gave 89Zr-DFO-NBA-onartuzumab in a crude RCC of 76% where purification by SEC gave a radiochemical purity of >99% and isolated radiochemical yield of 48%. Subsequent imaging and biodistribution studies showed high tumor uptake and specificity. After 72 h, tumor uptake of reached 13.8±1.9 %ID g−1 in the normal group and by comparison with blocking experiments tumor specificity was confirmed with ~44% tumor uptake reduction.

NBA provides a simple and fast method for the conjugation of 89Zr-DFO to native onartuzumab with favorable radiochemical conversions and yields compared with other light-activated handles like aryl azide3. We continue to explore the photochemical reactivity of NBA, and other functional groups, to access new classes of bioconjugate bonds.

 

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Figure 1 Cartoon depiction of the light-activated conjugation of 89Zr-DFO-NBA to the targeting mAb onartuzumab.

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  2. [2] Gherardi, E.; Birchmeier, W.; Birchmeier, C.; Woude, G. V. Nat. Rev. Cancer 2012, 12 (2), 89–103.
  3. [3] Klingler, S.; Fay, R.; Holland, J. P. J. Nucl. Med. 2020, 61 (7), 1072–1078.