Tetrazine-Based Cycloadditions: Application to Pretargeted Live Cell Imaging

Devaraj, Neal K.; Weissleder, Ralph; Hilderbrand, Scott A.

doi:10.1021/bc8004446

Cited by 763 publications

(694 citation statements)

References 24 publications

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“…It was found that the reactive dienes trans-cyclooctene 101 and norbornene 102 react relatively rapidly with suitable tetrazine dienophiles (which release nitrogen irreversibly on subsequent retro-[4 þ 2]-cycloaddition) allowing protein labelling at rates up to 1,000 times faster than SPAAC in the case of trans-cyclooctene (Fig. 4b).…”

Section: Review Nature Communications | Doi: 101038/ncomms5740mentioning

confidence: 99%

Selective chemical protein modification

2014

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Section: Review Nature Communications | Doi: 101038/ncomms5740mentioning

confidence: 99%

Selective chemical protein modification

2014

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“…Recent findings have established a set of bioorthogonal click reactions that do not require the cytotoxic copper catalyst used in early reports. These copper-free chemistries include strain-promoted azide-alkyne cycloaddition (SPAAC) and the inverse electron demand Diels-Alder reaction between tetrazine and norbornene [24,25]. Previous reports have used these click reactions primarily to crosslink click endfunctionalized branched polyethylene glycol (PEG) with linear crosslinkers composed of either PEG or linear peptides terminated with the appropriate click reaction pair [26][27][28][29].…”

Section: Introductionmentioning

confidence: 99%

Versatile click alginate hydrogels crosslinked via tetrazine–norbornene chemistry

et al. 2015

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Abstract:Alginate hydrogels are well-characterized, biologically inert materials that are used in many biomedical applications for the delivery of drugs, proteins, and cells. Unfortunately, canonical covalently crosslinked alginate hydrogels are formed using chemical strategies that can be biologically harmful due to their lack of chemoselectivity. In this work we introduce tetrazine and norbornene groups to alginate polymer chains and subsequently form covalently crosslinked click alginate hydrogels capable of encapsulating cells without damaging them. The rapid, bioorthogonal, and specific click reaction is irreversible and allows for easy incorporation of cells with high post-encapsulation viability. The swelling and mechanical properties of the click alginate hydrogel can be tuned via the total polymer concentration and the stoichiometric ratio of the complementary click functional groups. The click alginate hydrogel can be modified after gelation to display cell adhesion peptides for 2D cell culture using thiol-ene chemistry.Furthermore, click alginate hydrogels are minimally inflammatory, maintain structural integrity over several months, and reject cell infiltration when injected subcutaneously in mice. Click alginate hydrogels combine the numerous benefits of alginate hydrogels with powerful bioorthogonal click chemistry for use in tissue engineering applications involving the stable encapsulation or delivery of cells or bioactive molecules.

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“…However, many of these derivatives have low water solubility, require complex multistep synthesis, and possess suboptimal kinetics. Our search for alternative rapid, selective, and chemically accessible coupling reactions without need for a catalyst led us and others to investigate the [4 þ 2] inverse Diels-Alder cycloaddition (10)(11)(12)(13). We realized that this set of chemistries is more uniquely suited to biological applications and may indeed represent a universal platform technology ( Fig.…”

mentioning

confidence: 99%

Reactive polymer enables efficient in vivo bioorthogonal chemistry

Devaraj

Thurber

Keliher

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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There has been intense interest in the development of selective bioorthogonal reactions or "click" chemistry that can proceed in live animals. Until now however, most reactions still require vast surpluses of reactants because of steep temporal and spatial concentration gradients. Using computational modeling and design of pharmacokinetically optimized reactants, we have developed a predictable method for efficient in vivo click reactions. Specifically, we show that polymer modified tetrazines (PMT) are a key enabler for in vivo bioorthogonal chemistry based on the very fast and catalyst-free [4 þ 2] tetrazine/trans-cyclooctene cycloaddition. Using fluorescent PMT for cellular resolution and 18 F labeled PMT for whole animal imaging, we show that cancer cell epitopes can be easily reacted in vivo. This generic strategy should help guide the design of future chemistries and find widespread use for different in vivo bioorthogonal applications, particularly in the biomedical sciences.in vivo chemistry | pharmacokinetics | PET imaging | intravital microscopy | pretargeting T he ability to perform selective chemistries in living systems such as single cells, 3D cultures, invertebrates, or mammals would have far reaching applications in tracking biomolecules, designing new therapeutic approaches, and in visualizing medically relevant biomarkers. To date, only a few practical bioorthogonal reactions have been reported, the most popular being the Staudinger ligation and the [3 þ 2] cycloaddition "click" reaction between azides and alkynes (1, 2). The latter click reaction involves copper(I) catalyzed coupling of an azide and terminal alkyne to generate a stable triazole (2). Until recently, the necessity of the copper catalyst precluded use of this reaction in biological systems due to toxicity concerns (3, 4). Bertozzi and others elegantly solved this problem by developing several new ring strained dienophile derivatives that do not require catalysts (5-9). However, many of these derivatives have low water solubility, require complex multistep synthesis, and possess suboptimal kinetics. Our search for alternative rapid, selective, and chemically accessible coupling reactions without need for a catalyst led us and others to investigate the [4 þ 2] inverse Diels-Alder cycloaddition (10-13). We realized that this set of chemistries is more uniquely suited to biological applications and may indeed represent a universal platform technology ( Fig. 1 A and B). Specifically we and others have shown that the cycloaddition between tetrazine (Tz) and trans-cyclooctene (TCO) can proceed orders of magnitude faster than previously studied azide and alkyne click reactions and importantly does not require the action of a catalyst. This reaction has also been adapted for single cell imaging using newer fluorogenic Tz probes (14).Although initial work focused on in vitro labeling there has been a surge of recent work applying this reaction for various in vivo applications (15-17). Despite the above progress, our results with initial TCO...

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Tetrazine-Based Cycloadditions: Application to Pretargeted Live Cell Imaging

Cited by 763 publications

References 24 publications

Selective chemical protein modification

Selective chemical protein modification

Versatile click alginate hydrogels crosslinked via tetrazine–norbornene chemistry

Reactive polymer enables efficient in vivo bioorthogonal chemistry

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