Role of structural factors of antitumour anthraquinone derivatives and analogues in the ability to undergo bioreductive activation by NADPH cytochrome P450 reductase. Implications for increasing the activity against sensitive and multidrug-resistant leukaemia HL60 cells Dorota Kostrzewa-Nowak a , Bohdan Bieg b , Mark J.I. Paine c , C. Roland Wolf c and Jolanta Tarasiuk a The aim of this study was to examine the role of structural factors of antitumour anthraquinone derivatives and analogues in the ability to undergo bioreductive activation by NADPH cytochrome P450 reductase (CPR) and determine the impact of this activation on increasing the activity especially with regard to multidrug resistance (MDR) tumour cells. It was found that at a high NADPH concentration (500 lmol/l), the anthracenedione agent – ametantrone, with a nonmodified quinone structure, was susceptible to undergo CPR-dependent reductive activation. In contrast, it was evidenced that compounds with modified quinone groupment (benzoperimidine BP1, anthrapyridone CO1 and pyrazolopyrimidoacridine PPAC2) did not undergo reductive activation by CPR. This suggests that the presence of a modified quinone function is the structural factor excluding reductive activation of antitumour anthraquinone derivatives and analogues by CPR. In the second part of the work, the ability of antitumour anthraquinone derivatives and analogues to inhibit the growth of the human promyelocytic-sensitive leukaemia HL60 cell line as well as its MDR sublines exhibiting two different phenotypes of MDR related to the overexpression of P-glycoprotein (HL60/VINC) or MRP1 (HL60/DOX) was studied in the presence of exogenously added CPR. A significant increase in the activity of ametantrone with a nonmodified quinone structure after its reductive conversion by CPR was observed against HL60 as well as HL60/VINC and HL60/DOX cells, whereas in the case of quinone-modified compounds (BP1, CO1 and PPAC2), the presence of the activation system had no effect on their activity against the examined sensitive and MDR tumour cells. Anti-Cancer Drugs 00:000–000  c 2011 Wolters Kluwer Health | Lippincott Williams & Wilkins. Anti-Cancer Drugs 2011, 00:000–000 Keywords: anthraquinone derivatives and analogues, HL60 human promyelocytic leukaemia, multidrug resistance, NADPH cytochrome P450 reductase, reactive metabolites, redox cycling a Department of Biochemistry, University of Szczecin, b Department of Physics, Maritime University, Szczecin, Poland and c Cancer Research UK Molecular Pharmacology Unit, Biomedical Research Centre, Ninewells Hospital and Medical School, Dundee, UK Correspondence to Jolanta Tarasiuk, Department of Biochemistry, University of Szczecin, 3a Felczaka St, 71-412 Szczecin, Poland Tel/fax: + 44 48 914 441 550; e-mail: tarasiuk@univ.szczecin.pl Present address: Mark J.I. Paine, Liverpool School of Tropical Medicine, Liverpool, UK. Received 3 August 2011 Revised form accepted 29 November 2011 Introduction The clinical usefulness of bioreductive antitumour drugs is limited by the occurrence of multidrug resistance (MDR) associated with the presence of membrane transporters (e.g. P-glycoprotein, MRP1) belonging to the ATP-binding cassette protein family [1–3]. These transporters are responsible for the active ATP-depen- dent efflux of drugs out of resistant cells resulting in the decreased intracellular accumulation insufficient to inhibit resistant cell proliferation [3,4]. There is an increasing body of evidence indicating that the reductive activation of antitumour drugs, for example, anthracyclines, mitomycin C, tirapazamine and indolo- quinones could result in the formation of reactive intermediates capable of alkylation or crosslinking bind- ing with DNA [5–10] and may lead to a significant increase in the cytotoxic activity of these compounds against tumour cells [7,10–13]. Furthermore, our previous results found for the anthracycline drug – doxorubicin (DOX) and the anthracenedione agent – mitoxantrone (MX) [14,15] suggest that the bioreductive activation of antitumour compounds belonging to the anthraquinone family could generate reactive intermediates able to irreversibly bind to DNA before being removed from resistant cells by MDR exporting pumps, and provide a possibility to restore the activity of these drugs against multidrug-resistant tumour cells. In addition, it has been evidenced that the amount of NADPH was an essential factor determining the route of DOX as well MX activation by NADPH cytochrome P450 reductase (CPR) [14,15]. Similar findings were also reported for tirapazamine activation by this enzyme [11,12,16]. Thus, it is proposed that NADPH could participate in forming a coupled interactive system and, as a consequence, constitutes a control point in drug activation by cellular Original article 1 0959-4973  c 2011 Wolters Kluwer Health | Lippincott Williams & Wilkins DOI: 10.1097/CAD.0b013e32834fcf4f CE: Satish ED: Asra Op: Sridhar CAD 200930: LWW_CAD_200930