Shedding light by cancer redox—human NAD(P)H:quinone oxidoreductase 1 activation of a cloaked fluorescent dye

Silvers, William; Payne, Alex S.; McCarley, Robin L.

doi:10.1039/c1cc14578a

Cited by 46 publications

(48 citation statements)

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“…[12] The introduction of latent fluorescent agents based on TML provided new applications of this compound for imaging studies. [13] More recently, several fluorescence TML-based quinone agents were introduced for detecting DTD in cancer [14] and as theranostics for imaging as well as therapeutic applications. [15] Although these agents provide novel tools for detecting enzyme activity, the limited depth of view of fluorescence imaging within in vivo tissues demands another class of agents that detect enzyme activity in deep tissues.…”

mentioning

confidence: 99%

Detection of DT‐diaphorase Enzyme with a ParaCEST MRI Contrast Agent

Daryaei

Jones

Pagel

2017

Chemistry A European J

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mentioning

confidence: 99%

Detection of DT‐diaphorase Enzyme with a ParaCEST MRI Contrast Agent

Daryaei

Jones

Pagel

2017

Chemistry A European J

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“…42, 43 Our group has shown that reductively-activated liposomes can have their contents delivered upon self-cleavage of the reduced quinone propionic acid headgroup of the fusogenic lipids composing the liposomes; similarly, such cleavage of the reduced quinone from cloaked probes (prodyes) results in fluorescent signaling of human NQO1 activity. 32, 42 Variants of the quinone propionic acid (QPA) trigger group have also been investigated for prodrugs, such as those based on mustard 44 and oxindoles. 45 …”

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Human NAD(P)H:Quinone Oxidoreductase Type I (hNQO1) Activation of Quinone Propionic Acid Trigger Groups

et al. 2012

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NAD(P)H:quinone oxidoreductase type I (NQO1) is a target enzyme for triggered delivery of drugs at inflamed tissue and tumor sites, particularly those that challenge traditional therapies. Prodrugs, macromolecules, and molecular assemblies possessing trigger groups that can be cleaved by environmental stimuli are vehicles with the potential to yield active drug only at prescribed sites. Furthermore, quinone propionic acids (QPAs) covalently attached to prodrugs or liposome surfaces can be removed by application of a reductive trigger stimulus, such as that from NQO1; their rates of reductive activation should be tunable via QPA structure. We explored in detail the recombinant human NAD(P)H:quinone oxidoreductase type I (rhNQO1)-catalyzed NADH reduction of a family of substituted QPAs and obtained high precision kinetic parameters. It is found that small changes in QPA structure—in particular, single atom and function group substitutions on the quinone ring at R1—lead to significant impacts on the Michaelis constant (Km), maximum velocity (Vmax), catalytic constant (kcat), and catalytic efficiency (kcat/Km). Molecular docking simulations demonstrate that alterations in QPA structure result in large changes in QPA alignment and placement with respect to the flavin isoalloxazine ring in the active site of rhNQO1; a qualitative relationship exists between the kinetic parameters and the depth of QPA penetration into the rhNQO1 active site. From a quantitative perspective, a very good correlation is observed between log(kcat/Km) and the molecular-docking-derived distance between flavin hydride donor site and quinone hydride acceptor site in the QPAs, an observation that is in agreement with developing theories. The comprehensive kinetic and molecular modeling knowledge obtained for the interaction of recombinant human NQO1 with the quinone propionic acid analogues provides insight into the design and implementation of the QPA trigger groups for drug delivery applications.

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“…13,27 The 33-fold fluorescence enhancement for NI versus Q 3 NI and very large Stokes shift of 116 nm ( λ max, abs = 374 nm, λ max, em = 490 nm) bode well for use of Q 3 NI as a multifunctional turn-on probe for sensing and imaging applications that utilize reductive stimuli capable of initiating removal of the reduced quinone group. 28–31 …”

Section: Resultsmentioning

confidence: 99%

“…Thus, the rapid increase in fluorescence for reduced Q 3 NI results from naphthalimide dequenching caused by reduction-initiated removal of the quinone propionic acid group by lactonization. Due to the unique quenching mechanism of the Q 3 NI turn-on probe sensor, pronounced fluorescence signal enhancement upon revealing the NI reporter and its large Stokes shift, and the known resistance of the quinone propionic acid trigger group to reduction by other biological species, 31 we investigated Q 3 NI as a sensor probe of human NAD(P)H:quinone oxidoreductase isozyme 1 (hNQO1) activity in real-time biological applications.…”

Section: Resultsmentioning

confidence: 99%

“…From the plot shown in Figure 3, we obtained K m = 3.86 ± 0.79 μ M, V max = 0.037 ± 0.002 μ mol min −1 mg · NQO1 −1 , k cat = 0.019 ± 0.001 s −1 , and k cat / K m = 4.94 ± 0.33 × 10 3 M −1 s −1 . Due to the presence of the non-bulky ethanolamine linker and the need of only a single activation step to reveal reporter fluorescence, the kinetic constants are significantly higher than those of other NQO1 activatable fluorophores, 31,34,35 thereby ensuring sufficient signal enhancement for fast detection of hNQO1 activity.…”

Section: Resultsmentioning

confidence: 99%

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Profluorogenic Reductase Substrate for Rapid, Selective, and Sensitive Visualization and Detection of Human Cancer Cells that Overexpress NQO1

Silvers¹,

Prasai²,

et al. 2012

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Achieving the vision of identifying and quantifying cancer-related events and targets for future personalized oncology is predicated on the existence of synthetically accessible and economically viable probe molecules fully able to report the presence of these events and targets in a rapid, and highly selective and sensitive fashion. Delineated here are the design and evaluation of a newly synthesized turn-on probe whose intense fluorescent reporter signature is revealed only through probe activation by a specific intracellular enzyme present in tumor cells of multiple origins. Quenching of molecular probe fluorescence is achieved through unique photo-induced electron transfer (PeT) between the naphthalimide dye reporter and a covalently attached, quinone-based enzyme substrate. Fluorescence of the reporter dye is turned on by rapid removal of the quinone quencher, an event that immediately occurs only after highly selective, two-electron reduction of the sterically and conformationally restricted quinone substrate by the cancer-associated human NAD(P)H:quinone oxidoreductase isozyme 1 (hNQO1). Successes of the approach include rapid differentiation of NQO1-expressing and non-expressing cancer cell lines via the unaided eye, flow cytometry, fluorescence imaging, and two-photon microscopy. The potential for use of the turn-on probe in longer-term cellular studies is indicated by its lack of influence on cell viability and its in vitro stability.

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Shedding light by cancer redox—human NAD(P)H:quinone oxidoreductase 1 activation of a cloaked fluorescent dye

Abstract: A new quinone propionic acid-cloaked rhodamine fluorophore has its fluorescence revealed (de-cloaked) upon activation by human NAD(P)H:quinone oxidoreductase 1 (hNQO1), an upregulated enzyme in cancer cells and tumors.

Cited by 46 publications

References 34 publications

Detection of DT‐diaphorase Enzyme with a ParaCEST MRI Contrast Agent

Detection of DT‐diaphorase Enzyme with a ParaCEST MRI Contrast Agent

Human NAD(P)H:Quinone Oxidoreductase Type I (hNQO1) Activation of Quinone Propionic Acid Trigger Groups

Profluorogenic Reductase Substrate for Rapid, Selective, and Sensitive Visualization and Detection of Human Cancer Cells that Overexpress NQO1

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