Mechanism‐Based Fluorogenic trans‐Cyclooctene–Tetrazine Cycloaddition

Abstract

The development of fluorogenic reactions which lead to the formation of fluorescent products from two nonfluorescent starting materials is highly desirable,b ut challenging.R eported herein is an ew concept of fluorescent product formation upon the inverse electron-demand Diels-Alder reaction of 1,2,4,5-tetrazines with particular trans-cyclooctene (TCO) isomers.Insharp contrast to knownfluorogenic reagents the presented chemistry leads to the rapid formation of unprecedented fluorescent 1,4-dihydropyridazines so that the fluorophore is built directly upon the chemical reaction. Attachment of an extra fluorophore moiety is therefore not needed. The photochemical properties of the resulting dyes can be easily tuned by changing the substitution pattern of the starting 1,2,4,5-tetrazine.W es upport the claim with NMR measurements and rationalizethe data by computational study. Cell-labeling experiments were performed to demonstrate the potential of the fluorogenic reaction for bioimaging.

An Extended Approach for the Development of Fluorogenic trans‐Cyclooctene–Tetrazine Cycloadditions

“…Indeed, during experimentation with various 1,2,4,5‐tetrazines we found that tetrazine derivative 1 a , bearing a p ‐dimethylamino substituent, forms a fluorescent product when combined with equatorially hydroxy‐substituted TCO 2 a (eqTCO). As we have previously reported, the same derivative leads to fluorescent products, with different photophysical properties, upon reaction with the corresponding axially hydroxy‐substituted TCO 2 b (axTCO, Figure B and C). We ascribed the formation of the two different fluorescent species to the production of different tautomers of the dihydropyridazine core (Figures S1 and S2 in the Supporting Information).…”

“…The first reaction step of inverse‐electron‐demand Diels–Alder (iEDDA) cycloaddition between a 1,2,4,5‐tetrazine and a dienophile involves the formation of a tetraazabicyclic system, which, after a retro‐Diels–Alder reaction and extrusion of molecular nitrogen, is transformed into a 4,5‐dihydropyridazine (Figure A). This can further isomerize to the corresponding 1,4‐dihydropyridazine through, for example, addition and elimination of a water molecule or alternatively through intramolecular interaction with an appropriately placed heteroatom substituent on the TCO . The initially formed 4,5‐dihydropyridazine heterocyclic core is a unique π‐conjugated system formed upon reaction between tetrazines and all TCO derivatives.…”

Fishing for Drug Targets: A Focus on Diazirine Photoaffinity Probe Synthesis

“…There are further developments in the tetrazine Diels−Alder reagents where these can be tuned according to pendent groups. 55 Unfortunately these initial cyclopropene photo-cross-linkers caused high nonspecific background labeling. 33 A further example of live-cell imaging was conducted with the most advanced clinical Aurora kinase cell cycle inhibitor, MLN8237.…”

Uncovering the key role of distortion in tetrazine ligations guides the design of bioorthogonal tools that defy the reactivity/stability tradeoff

“…[33][34][35] 4-Pyridyl-Tz have previously been used for the design of fluorogenic probes. [36] However, so far there is no comparative data on the IEDDA reaction kinetics of 2-pyridyl-and 4pyridyl-tetrazines. Therefore, we selected a series of phenyl (Ph), 2-pyridyl (2Pyr), 3-pyridyl (3Pyr), and 4-pyridyl (4Pyr) substituted Tz (Fig.…”