Background: The biotin-avidin interaction remains a gold standard for the two-step pretargeting approach to image tumor sites. We aim to develop two-step pretargeting systems utilizing (99m)Tc labeled biotin functionalized macrocyclic chelating agents synthesized using the highly efficient Cu(I) catalyzed azide-alkyne cycloaddition for potential radioimaging applications.
Methods: A facile synthesis of DOTA-Triazole-Biotin, radiocomplexation with (99m)Tc and the pretargeting protocol is described. The synthesis features Cu(I) catalyzed click conjugation between biotinylated azide and propynyl functionalized DO3A. (99m)Tc radiolabeling was performed to detect the accumulation of avidin as the pretargeting agent. Cytotoxicity was determined using the trypan blue exclusion assay, macrocolony, and MTT assay. Cell uptake studies were performed using radiolabeled DOTA-Triazole-Biotin and compared with avidin treated cells for 2 h. Tumor imaging was performed in U-87MG cell line implanted tumor bearing nude mice and uptake of the radiotracer was estimated.
Results: All compounds have been successfully characterized by NMR and MS spectroscopy. More than 96% radiolabeling efficiency was obtained and the radioconjugate exhibited sufficient stability under physiological conditions.
Conclusion: To summarize, a new candidate for avidin based two-step pretargeting of tumors has been synthesized and evaluated for potential imaging and diagnostic applications. The chelate possesses high stability under physiological conditions, exhibits effective interaction with its avidin target, and low nonspecific retention in vivo.