Synthetically Tractable Click Hydrogels for Three-Dimensional\nCell Culture Formed Using Tetrazine–Norbornene Chemistry

Abstract

The implementation of bio-orthogonal\nclick chemistries is a topic\nof growing importance in the field of biomaterials, as it is enabling\nthe development of increasingly complex hydrogel materials capable\nof providing dynamic, cell-instructive microenvironments. Here, we\nintroduce the tetrazine–norbornene inverse electron demand\nDiels–Alder reaction as a new cross-linking chemistry for the\nformation of cell laden hydrogels. The fast reaction rate and irreversible\nnature of this click reaction allowed for hydrogel formation within\nminutes when a multifunctional PEG-tetrazine macromer was reacted\nwith a dinorbornene peptide. In addition, the cytocompatibility of\nthe polymerization led to high postencapsulation viability of human\nmesenchymal stem cells, and the specificity of the tetrazine–norbornene\nreaction was exploited for sequential modification of the network\nvia thiol–ene photochemistry. These advantages, combined with\nthe synthetic accessibility of the tetrazine molecule compared to\nother bio-orthogonal click reagents, make this cross-linking chemistry\nan interesting and powerful new tool for the development of cell-instructive\nhydrogels for tissue engineering applications.