A major goal of personalized medicine in oncology is the identification of drugs with predictable efficacy based on a specific trait of the cancer cell, as has been demonstrated with gleevec (presence of Bcr-Abl protein), herceptin (Her2 overexpression), and iressa (presence of a specific EGFR mutation). This is a challenging task, as it requires identifying a cellular component that is altered in cancer, but not normal cells, and discovering a compound that specifically interacts with it. The enzyme NQO1 is a potential target for personalized medicine, as it is overexpressed in many solid tumors. In normal cells NQO1 is inducibly expressed, and its major role is to detoxify quinones via bioreduction; however, certain quinones become more toxic after reduction by NQO1, and these compounds have potential as selective anticancer agents. Several quinones of this type have been reported, including mitomycin C, RH1, EO9, streptonigrin, β-lapachone, and deoxynyboquinone (DNQ). However, no unified picture has emerged from these studies, and the key question regarding the relationship between NQO1 processing and anticancer activity remains unanswered. Here, we directly compare these quinones as substrates for NQO1 in vitro, and for their ability to kill cancer cells in culture in an NQO1-dependent manner. We show that DNQ is a superior NQO1 substrate, and we use computationally guided design to create DNQ analogues that have a spectrum of activities with NQO1. Assessment of these compounds definitively establishes a strong relationship between in vitro NQO1 processing and induction of cancer cell death and suggests these compounds are outstanding candidates for selective anticancer therapy.

Efficient NQO1 substrates are potent and selective anticancer agents.

Our previous study showed that the methanolic extract of Inula helenium (elecampane) had the potential to induce detoxifying enzymes such as quinine reductase (QR) and glutathione S-transferase. In this study we further fractionated the methanolic extract into hexane-, dichloromethane-, butanol-, and water-soluble fractions according to polarity. The hexane fraction showed the highest QR-inducing activity and also induced glutathione S-transferase in a dose-dependent manner. Its potential to induce the reporter activity suggested an antioxidant response element-mediated mechanism of action in the induction of phase II detoxifying enzymes. Intraperitoneal injection of the hexane fraction of I. helenium into ICR mice caused a significant increase of QR activity in liver, kidney, small intestine, and stomach. Sesquiterpenes, isolated from the hexane fraction, appeared to be major components responsible for QR induction. Among the seven compounds tested in this study, alantolactone, isoalantolactone, and 5α-e...

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Induction of Detoxifying Enzyme by Sesquiterpenes Present in Inula helenium

2,5-Diaziridinyl-3-(hydroxymethyl)-6-methyl-1,4-benzoquinone (RH1) is a novel antitumor diaziridinyl benzoquinone derivative designed to be bioactivated by the two-electron reductase NAD(P)H:quinone oxidoreductase (NQO1) and is currently in clinical trials. NQO1 is expressed at high levels in many solid tumors. RH1 cytotoxicity has been shown previously to be NQO1-dependent. The purpose of this study was to investigate whether other reducing enzymes such as cytochrome b(5) reductase (b5R), cytochrome P450 reductase (P450R), dihydronicotinamide riboside:quinone oxidoreductase 2 (NQO2), and xanthine oxidase/xanthine dehydrogenase (XO/XDH) also contribute to the bioactivation and cytotoxicity of RH1 in human tumor cells. For these studies, we established a series of stable MDA468 breast cancer cell lines overexpressing various levels of NQO1, b5R, P450R, and NQO2 and compared RH1-induced growth inhibition [3-(4,5-dimethylthiazol-2,5-diphenyl)tetrazolium and sulforhodamine B analysis] and interstrand DNA cross-linking (comet analysis) in both parental MDA468 cells and transfected clones. RH1 toxicity correlated with NQO1 and NQO2 but not with either b5R or P450R activity levels in the respective series of transfected MDA468 cell clones. Enzymatic assays showed that RH1 was an in vitro substrate for xanthine oxidase. However, XO/XDH protein and activity could not be detected in a variety of human tumor cell lines. These studies suggest that NQO1 and NQO2 are the principal enzymatic determinants of RH1 bioactivation in MDA468 tumor cells and that b5R, P450R, and XDH/XO are unlikely to play major roles. Our studies also suggest that NQO2 may be particularly relevant as a bioactivation system for RH1 in NQO1-deficient tumors such as leukemias and lymphomas.

Dissecting the role of multiple reductases in bioactivation and cytotoxicity of the antitumor agent 2,5-diaziridinyl-3-(hydroxymethyl)-6-methyl-1,4-benzoquinone (RH1).

Herpesvirus infections are a leading cause of neurodevelopmental delay in newborns and end-organ disease in immunocompromised patients. One leading strategy to reduce the disease burden of herpesvirus infections such as herpes simplex virus (HSV) and human cytomegalovirus (HCMV) is to prevent primary acquisition by vaccination, yet vaccine development remains hampered by limited understanding of immune correlates of protection against infection. Traditionally, vaccine development has aimed to increase antibody titers with neutralizing function, which involves the direct binding of antibodies to viral particles. However, recent research has explored the numerous other responses that can be mediated by engagement of the antibody constant region (Fc) with Fc receptors (FcR) present on immune cells or with complement molecules. These functions include antiviral responses such as antibody-dependent cell-mediated cytotoxicity (ADCC) and antibody-dependent cellular phagocytosis (ADCP). Uniquely, herpesviruses encode FcR that can act as distractor receptors for host antiviral IgG, thus enabling viral evasion of host defenses. This review focuses on the relative roles of neutralizing and non-neutralizing functions antibodies that target herpesvirus antigens for HSV and HCMV, as well as the roles of Fc-FcR interactions for both host defenses and viral escape.

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The Roles of Host and Viral Antibody Fc Receptors in Herpes Simplex Virus (HSV) and Human Cytomegalovirus (HCMV) Infections and Immunity.

Phase I pharmacokinetic and pharmacodynamic study of the bioreductive drug RH1

The elevated expression of the flavoprotein NAD(P)H:quinone acceptor oxidoreductase (NQO1) (EC 1.6.99.2) in many human solid tumors, along with its ability to activate quinone-based anticancer agents, makes it an excellent target for enzyme-directed drug development. Previous studies have shown a significant statistical correlation between NQO1 enzymatic activity and the cytotoxicity of certain antitumor quinones. RH1 [2,5-diaziridinyl-3-(hydroxymethyl)-6-methyl-1,4-benzoquinone], presently in late preclinical and entering early clinical development, has been previously considered to be an excellent substrate for activation by NQO1. In this study we investigate the cytotoxicity of RH1 in cell lines selected from the NCI's 60 tumor cell line panel, expressing varying levels of NQO1 activity. Exposure time- and concentration-dependent cytotoxicity was seen, apparently independent from levels of NQO1 activity in these cells. Furthermore, the NQO1 inhibitor dicoumarol had no impact on the sensitivity profiles of RH1 response. The HL-60 myeloid leukemia cells, which do not have detectable NQO1 activity, were further investigated. RH1 treatment of HL-60 cells generated high levels of free radicals, which was accompanied by robust redox cycling, oxygen consumption and induction of apoptosis. These results are in agreement with previous data suggesting that, in addition to its activation by NQO1, RH1-induced cytotoxicity might involve alternative pathways for activation of this compound. Furthermore, the high cytotoxicity of RH1 in the leukemia/lymphoma subpanel of the NCI in vitro cell line screen would suggest an empirical rationale for the utilization of this compound in the treatment of these malignancies.

Christopher M Nelson

Papers

Cytotoxicity of RH1: NAD(P)H:quinone acceptor oxidoreductase (NQO1)-independent oxidative stress and apoptosis induction.