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An improved cell-permeable fluorogenic substrate as the basis for a highly sensitive test for NAD(P)H quinone oxidoreductase 1 (NQO1) in living cells

Cuff, Simone, Lewis, Ruth, Chinje, Edwin, Jaffar, Mohammed, Knox, Richard and Weeks, Ian 2018. An improved cell-permeable fluorogenic substrate as the basis for a highly sensitive test for NAD(P)H quinone oxidoreductase 1 (NQO1) in living cells. Free Radical Biology and Medicine 116 , pp. 141-148. 10.1016/j.freeradbiomed.2018.01.009

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Abstract

NAD(P)H:quinone oxidoreductase 1 (NQO1) is a flavoenzyme upregulated in response to oxidative stress and in some cancers. Its upregulation by compounds has been used as an indicator of their potential anti-cancer properties. In this study we have designed, produced and tested a fluorogenic coumarin conjugate which selectively releases highly fluorescent 4-methylumbelliferone (4-MU) in the presence of NQO1. It was found that measuring 4-MU release rapidly and specifically quantitated NQO1 levels in vitro and in live cells. Both the substrate and its products freely perfused through cell membranes and were non-toxic. The substrate was very specific with low background, and the assay itself could be done in less than 10 minutes. This is the first assay to allow the quantitation of NQO1 in live cells which can then be retained for further experiments.

Item Type: Article
Date Type: Publication
Status: Published
Schools: Medicine
Publisher: Elsevier
ISSN: 0891-5849
Date of Acceptance: 6 January 2018
Last Modified: 31 Mar 2018 20:53
URI: http://orca.cf.ac.uk/id/eprint/108854

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