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Bohrmann, Johannes Lennart

University of British Columbia

The bioorthogonal click reaction between tetrazines and trans-cyclooctene (TCO) has tremendous potential for bioconjugation and pretargeted imaging due to its rapid kinetics and selectivity. The aim of this dissertation was to utilize the tetrazine ligation for the synthesis of an aptamer targeted nanoprobe and to evaluate novel 99mTc-labeled tetrazines for pretargeted imaging. Aptamers are small single-stranded oligonucleotides that bind their target with high specificity and affinity. A significant limitation of aptamers is their small size and susceptibility to nucleases, which leads to rapid degradation and elimination. Formulation strategies such as the conjugation to macromolecules or nanoparticles hold great potential to improve the pharmacokinetics of aptamers. In this dissertation, the tetrazine ligation was used to covalently conjugate sgc8, a PTK7 specific DNA aptamer to the long circulating polymeric nanoparticle hyperbranched polyglycerol (HPG). The conjugation of sgc8 to HPG led to a marked increase in plasma stability. The binding ability of sgc8 and HPG-sgc8 was confirmed against a range of cancer derived cell lines. However, control experiments with a non-targeting aptamer under different temperatures revealed important limitations regarding the specificity of sgc8 and non-specific binding of ssDNA. The pharmacokinetic profile and tumor uptake of targeted and non-targeted HPG was assessed using SPECT/CT imaging. Despite strong binding in vitro, no targeting effect of HPG-sgc8 could be detected in vivo. This study suggests that passive accumulation of HPG in the tumor dominates over sgc8-mediated binding and highlights the need to better characterize existing aptamers. Pretargeted imaging with radiolabeled tetrazines holds the potential to reduce radiation burden to patients and improve contrast. Structure-activity relationships for many radiolabeled tetrazines have been established over the recent years. However, a lack of knowledge exists concerning ⁹⁹ᵐTc-labeled tetrazines. A panel of four ⁹⁹ᵐTc-labeled was synthesized and the in vivo click performance was assessed using a TCO-modified bisphosphonate for bone targeting. The results of this study indicate that less hydrophilic tetrazines and longer plasma half-life might be important characteristics to further develop ⁹⁹ᵐTc-labeled tetrazines.

Text

2024-09-30

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/86043

Pharmaceutical Sciences

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/