Development of Bioorthogonal Chemistry and Its Applications

This thesis traces the evolution of bioorthogonal chemistry from its conceptual origins to its pubblic recognition by the Nobel Prize. The journey begins with Sharpless's click chemistry paradigm, which established criteria for ideal reactions, exemplified by Copper Click Chemistry, though with copper toxicity limitation in living systems. Bertozzi's Staudinger Ligation introduced the first truly bioorthogonal reaction, demonstrating that abiotic functional groups could operate selectively within cells. Strain-promoted azide-alkyne cycloaddition followed, replacing toxic catalysts with ring strain as a driving force, later optimized with ring modifications to further exploit the potentials. The inverse electron-demand Diels-Alder (IEDDA) Tetrazine Ligation with trans-cyclooctene synthesized these principles, combining maximal strain energy with inverse electron-demand control to achieve exceptional kinetics. Substituent effects enable electronic tuning of reactivity, while pH-sensitive variants allow targeted applications in biological environments. The Click-to-Release strategy further transforms simple ligation into triggerable drug delivery. The tetrazine-TCO reaction stands as the culmination of bioorthogonal design principles and a foundation for future clinical applications.