Abstract/Summary

This project introduces novel targeting approaches to increase the selectivity of chemotherapies in cancer treatment. These approaches involve converting the chemotherapies into less cytotoxic prodrugs that can be subsequently activated by bioorthogonal reactions to restore their cytotoxicity at the desired site of action. Two triphenylphosphine-prodrugs (triphenylphosphine-doxorubicin and triphenylphosphine-N-mustard, yielded in 39% and 31%, and with 95% and 98% purity, respectively) and two trans-cyclooctene-prodrugs (TCO-doxorubicin and TCO-N-mustard, yielded in 50% and 45%, and with 97% and 96% purity, respectively) were synthesised and tested for activation by azide and tetrazine activators via the Staudinger ligation and tetrazine ligation reactions, respectively. Two complementary approaches were used for the selective delivery of the activators to tumour cells. In the first, metabolic glycoengineering was used to intercept the biosynthesis of overexpressed tumour associated carbohydrate antigens within cancer cells’ surface glycans. Selective installation of azide and tetrazine tags on breast cancer cells using azide (50 µM)- and tetrazine (10 and 20 µM)- modified monosaccharides was successfully achieved (~17-fold and 35-65 folds higher than on non-cancerous fibroblasts). Subsequent activation of the triphenylphosphine- and TCO-prodrugs (> 68 % and up to 100%, respectively) by the engineered azides and tetrazines was demonstrated. In the second approach, PEG activators (PEG-azide and PEG-tetrazine, 10 KDa) were used for targeting cancer via passive targeting and the feasibility of their utilisation for prodrug activation was investigated. HPLC-monitored release studies showed 100% release of doxorubicin from the triphenylphosphine-doxorubicin and the TCO-doxorubicin prodrugs after 24 hours and 20 minutes by PEG-azide and PEG-tetrazine, respectively. In vitro activation studies on breast cancer cells showed the activation of the triphenylphosphine- and TCO-prodrugs (~68-76% and 100%, respectively) by PEG-azide and PEG-tetrazine. Taken together, these resultsshow the feasibility of utilising metabolic glycoengineering and passive targeting as targeting approaches to increase the selectivity of bioorthogonal prodrugs activation in breast cancer.