Fluorescent probest hat light-up upon reaction with complementary bioorthogonal reagents are superior tools for no-wash fluorogenic bioimaging applications.I n this work, at horough study is presented on as et of seventeen structurally diverse coumarin-tetrazine probes that produce fluorescent dyes with exceptional turn-onr atios when reacted with trans-cyclooctene (TCO) and bicyclononyne (BCN) dienophiles. In general,f ormation of the fully aromatic pyridazine-containing dyes resulting fromt he reaction with BCN was found superiori nt erms of fluorogenicity.H owever, evaluation of the probesi nc ellular imaging experiments revealed that other factors, such as reaction kinetics and good cell permeability,p revail over the fluorescence turn-on properties. The best compound identified in this study showed excellent performance in live cell-labeling experimentsa nd enabledn o-wash fluorogenic imaging on at imescale of seconds.

Photochemical generation of dibenzosilacyclohept-4-yne 3 from the corresponding cyclopropenone 1 and its copper-free click reactions are reported. Steady-state irradiation, kinetic, and transient absorption spectroscopy studies revealed that strained alkyne 3 is rapidly (<5 ns) and efficiently (Φ = 0.58-0.71) photoreleased from 1 and undergoes remarkably fast, selective, and high-yielding 1,3-dipolar cycloaddition with benzyl azide (∼20 M s) or [4 + 2] inverse-electron-demand Diels-Alder reaction with 1,2,4,5-tetrazines (∼260 M s) in both methanol and acetonitrile.

Mitragynine (MG) is the most abundant alkaloid component of the psychoactive plant material “kratom”, which according to numerous anecdotal reports shows efficacy in self-medication for pain syndromes, depression, anxiety, and substance use disorders. We have developed a synthetic method for selective functionalization of the unexplored C11 position of the MG scaffold (C6 position in indole numbering) via the use of an indole-ethylene glycol adduct and subsequent iridium-catalyzed borylation. Through this work we discover that C11 represents a key locant for fine-tuning opioid receptor signaling efficacy. 7-Hydroxymitragynine (7OH), the parent compound with low efficacy on par with buprenorphine, is transformed to an even lower efficacy agonist by introducing a fluorine substituent in this position (11-F-7OH), as demonstrated in vitro at both mouse and human mu opioid receptors (mMOR/hMOR) and in vivo in mouse analgesia tests. Low efficacy opioid agonists are of high interest as candidates for generating safer opioid medications with mitigated adverse effects.

The development of abiotic chemical reactions that can be performed in an organelle-specific manner can provide new opportunities in drug delivery and cell and chemical biology. However, due to the complexity of the cellular environment, this remains a significant challenge. Here, we introduce structurally redesigned bioorthogonal tetrazine reagents that spontaneously accumulate in mitochondria of live mammalian cells. The attributes leading to their efficient accumulation in the organelle were optimized to include the right combination of lipophilicity and positive delocalized charge. The best performing mitochondriotropic tetrazines enable subcellular chemical release of TCO-caged compounds as we show using fluorogenic substrates and mitochondrial uncoupler niclosamide. Our work demonstrates that a shrewd redesign of common bioorthogonal reagents can lead to their transformation into organelle-specific probes, opening the possibility to activate prodrugs and manipulate biological processes at the subcellular level by using purely chemical tools.