NAD(P)H:Quinone Oxidoreductase-1-Dependent and -Independent Cytotoxicity of Potent Quinone Cdc25 Phosphatase Inhibitors

Abstract

Cdc25 dual-specificity phosphatases coordinate cell cycle progression and cellular signaling. Consequently, Cdc25 inhibitors represent potential anticancer agents. We evaluated Ͼ10,000 compounds for inhibition of human Cdc25 phosphatases and identified many potent and selective inhibitors, which all contained a quinone. Bioreductive enzymes frequently detoxify or activate quinones. Therefore, we evaluated the effect of NAD(P)H:quinone oxidoreductase-1 (NQO1) and reductase-rich microsomes on the activity of three quinone-containing Cdc25 inhibitors: 2-(2-hydroxyethylsulfanyl)-3-methyl-1,4-naphthoquinone (Cpd 5, compound 5; NSC 672121), 2,3-bis-(2-hydroxyethylsulfanyl)-1,4-naphthoquinone (NSC 95397), and 6-chloro-7-(2-morpholin-4-ylethylamino)quinoline-5,8-dione (NSC 663284). Each inhibitor was reduced by human NQO1 (K m of 0.3-0.5 M) but none by microsomes. Compounds were evaluated with six cancer cell lines containing different amounts of NQO1: HT-29 (1056 nmol/mg/ min), HCT116 (660 nmol/mg/min), sublines HCT116-R30A (28 nmol/mg/min) and HCT-116R30A/NQ5 (934 nmol/mg/min), MDA-MB-231/Q2 (null NQO1), and subline MDA-MB-231/Q6 (124 nmol/mg/min) but containing similar amounts of microsomal cytochrome P450 reductase and cytochrome b 5 reductase. Growth inhibition and G2/M arrest by Cpd 5 was proportional to NQO1 levels, requiring 4-to 5-fold more Cpd 5 to inhibit HCT-116 or HCT-116R30A/NQ5 compared with HCT-116R30A. In contrast, in all tested cell lines irrespective of NQO1 level, growth inhibition and G2/M arrest by NSC 95375 and NSC 663284 were similar (average IC 50 of 1.3 Ϯ 0.3 and 2.6 Ϯ 0.4 M, respectively). NSC 95375 and NSC 663284 also caused similar Cdk1 hyperphosphorylation, indicating similar Cdc25 inhibition. However, lower Cpd 5 concentrations were needed to produce Cdk1 hyperphosphorylation in sublines with minimal NQO1. Thus, NQO1 detoxified Cpd 5, probably by reducing it to a less active hydroquinone, whereas NSC 95397-and NSC 663284-generated cytotoxicity was unaffected by NQO1.

Chemical synthesis and biological evaluation of a NAD(P)H:quinone oxidoreductase-1–targeted tripartite quinone drug delivery system

“…The results of this study show that the prototype prodrug 1 is an efficient substrate for purified human recombinant NQO1 with apparent V max (11.86 F 3.09 Amol/min/mg) and K m (2.70 F 1.14 Amol/L) values that are similar to other quinone-based bioreductive drugs such as EO9 (8,28). The affinity of NQO1 for this prodrug is 10 times lower than its affinity for its best known substrate, menadione, which has a K m of 28 Amol/L and a V max of 305 Amol/min/mg (34). It should be stated however that this analysis is complicated by the fact that prodrug 1 degrades to spirolactone 7 in aqueous conditions (35).…”

Redox Regulation of Cdc25B by Cell-Active Quinolinediones

“…8). Although DA3003-1 seemed to participate in redox cycling in cells, cell lines differing in levels of NAD(P)H:quinone oxidoreductase-1 are equally sensitive to DA3003-1 cytotoxic effects (Han et al, 2004). Therefore, these compounds may be substrates for other quinone reductases or dehydrogenases, such as carbonyl reductase and xanthine dehydrogenase.…”

PM-20, a novel inhibitor of Cdc25A, induces extracellular signal–regulated kinase 1/2 phosphorylation and inhibits hepatocellular carcinoma growth in vitro and in vivo

“…An important feature is that the maleimide derivatives are potent Michael acceptors and are expected to react covalently with protein tyrosine phosphatase active site cysteines. However, unlike the quinone derivatives, PM-20 is not able to undergo redox cycling, which is thought to limit the activity of the former because the reduced hydroquinone form is incapable of undergoing the Michael reaction (32).…”