Sarah R Klein

U.S. Food and Drug Administration, Washington, D. C., DC, United States

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Publications (4)20.75 Total impact

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    ABSTRACT: The iron chelator Dp44mT is a potent topoisomerase II╬▒ inhibitor with novel anticancer activity. Doxorubicin (Dox), the current front-line therapy for breast cancer, induces a dose-limiting cardiotoxicity, in part through an iron-mediated pathway. We tested the hypothesis that Dp44mT can improve clinical outcomes of treatment with Dox by alleviating cardiotoxicity. The general cardiac and renal toxicities induced by Dox were investigated in the presence and absence of Dp44mT. The iron chelating cardioprotectant Dexrazoxane (Drz), which is approved for this indication, was used as a positive control. In vitro studies were carried out with H9c2 rat cardiomyocytes and in vivo studies were performed using spontaneously hypertensive rats. Testing of the GI(50) profile of Dp44mT in the NCI-60 panel confirmed activity against breast cancer cells. An acute, toxic dose of Dox caused the predicted cellular and cardiac toxicities, such as cell death and DNA damage in vitro and elevated cardiac troponin T levels, tissue damage, and apoptosis in vivo. Dp44mT alone caused insignificant changes in hematological and biochemical indices in rats, indicating that Dp44mT is not significantly cardiotoxic as a single agent. In contrast to Drz, Dp44mT failed to mitigate Dox-induced cardiotoxicity in vivo. We conclude that although Dp44mT is a potent iron chelator, it is unlikely to be an appropriate cardioprotectant against Dox-induced toxicity. However, it should continue to be evaluated as a potential anticancer agent as it has a novel mechanism for inhibiting the growth of a broad range of malignant cell types while exhibiting very low intrinsic toxicity to healthy tissues.
    Cancer Chemotherapy and Pharmacology 03/2011; 68(5):1125-34. · 2.80 Impact Factor
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    ABSTRACT: Mitoquinone (MitoQ) is a synthetically modified, redox-active ubiquinone compound that accumulates predominantly in mitochondria. We found that MitoQ is 30-fold more cytotoxic to breast cancer cells than to healthy mammary cells. MitoQ treatment led to irreversible inhibition of clonogenic growth of breast cancer cells through a combination of autophagy and apoptotic cell death mechanisms. Relatively limited cytotoxicity was seen with the parent ubiquinone coenzyme Q(10.) Inhibition of cancer cell growth by MitoQ was associated with G(1)/S cell cycle arrest and phosphorylation of the checkpoint kinases Chk1 and Chk2. The possible role of oxidative stress in MitoQ activity was investigated by measuring the products of hydroethidine oxidation. Increases in ethidium and dihydroethidium levels, markers of one-electron oxidation of hydroethidine, were observed at cytotoxic concentrations of MitoQ. Keap1, an oxidative stress sensor protein that regulates the antioxidant transcription factor Nrf2, underwent oxidation, degradation, and dissociation from Nrf2 in MitoQ-treated cells. Nrf2 protein levels, nuclear localization, and transcriptional activity also increased following MitoQ treatment. Knockdown of Nrf2 caused a 2-fold increase in autophagy and an increase in G(1) cell cycle arrest in response to MitoQ but had no apparent effect on apoptosis. The Nrf2-regulated enzyme NQO1 is partly responsible for controlling the level of autophagy. Keap1 and Nrf2 act as redox sensors for oxidative perturbations that lead to autophagy. MitoQ and similar compounds should be further evaluated for novel anticancer activity.
    Journal of Biological Chemistry 11/2010; 285(45):34447-59. · 4.65 Impact Factor
  • [show abstract] [hide abstract]
    ABSTRACT: Mitoquinone (MitoQ) is a synthetically modified, redox-active ubiquinone compound that accumulates predominantly in mitochondria. We found that MitoQ is 30-fold more cytotoxic to breast cancer cells than to healthy mammary cells. MitoQ treatment led to irreversible inhibition of clonogenic growth of breast cancer cells through a combination of autophagy and apoptotic cell death mechanisms. Relatively limited cytotoxicity was seen with the parent ubiquinone coenzyme Q10. Inhibition of cancer cell growth by MitoQ was associated with G1/S cell cycle arrest and phosphorylation of the checkpoint kinases Chk1 and Chk2. The possible role of oxidative stress in MitoQ activity was investigated by measuring the products of hydroethidine oxidation. Increases in ethidium and dihydroethidium levels, markers of one-electron oxidation of hydroethidine, were observed at cytotoxic concentrations of MitoQ. Keap1, an oxidative stress sensor protein that regulates the antioxidant transcription factor Nrf2, underwent oxidation, degradation, and dissociation from Nrf2 in MitoQ-treated cells. Nrf2 protein levels, nuclear localization, and transcriptional activity also increased following MitoQ treatment. Knockdown of Nrf2 caused a 2-fold increase in autophagy and an increase in G1 cell cycle arrest in response to MitoQ but had no apparent effect on apoptosis. The Nrf2-regulated enzyme NQO1 is partly responsible for controlling the level of autophagy. Keap1 and Nrf2 act as redox sensors for oxidative perturbations that lead to autophagy. MitoQ and similar compounds should be further evaluated for novel anticancer activity.
    Journal of Biological Chemistry 11/2010; 285(45):34447-34459. · 4.65 Impact Factor
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    ABSTRACT: Di-2-pyridylketone-4,4,-dimethyl-3-thiosemicarbazone (Dp44mT) is being developed as an iron chelator with selective anticancer activity. We investigated the mechanism whereby Dp44mT kills breast cancer cells, both as a single agent and in combination with doxorubicin. Dp44mT alone induced selective cell killing in the breast cancer cell line MDA-MB-231 when compared with healthy mammary epithelial cells (MCF-12A). It induces G(1) cell cycle arrest and reduces cancer cell clonogenic growth at nanomolar concentrations. Dp44mT, but not the iron chelator desferal, induces DNA double-strand breaks quantified as S139 phosphorylated histone foci (gamma-H2AX) and Comet tails induced in MDA-MB-231 cells. Doxorubicin-induced cytotoxicity and DNA damage were both enhanced significantly in the presence of low concentrations of Dp44mT. The chelator caused selective poisoning of DNA topoisomerase IIalpha (top2alpha) as measured by an in vitro DNA cleavage assay and cellular topoisomerase-DNA complex formation. Heterozygous Nalm-6 top2alpha knockout cells (top2alpha(+/-)) were partially resistant to Dp44mT-induced cytotoxicity compared with isogenic top2alpha(+/+) or top2beta(-/-) cells. Specificity for top2alpha was confirmed using top2alpha and top2beta small interfering RNA knockdown in HeLa cells. The results show that Dp44mT is cytotoxic to breast cancer cells, at least in part, due to selective inhibition of top2alpha. Thus, Dp44mT may serve as a mechanistically unique treatment for cancer due to its dual ability to chelate iron and inhibit top2alpha activity.
    Cancer Research 02/2009; 69(3):948-57. · 8.65 Impact Factor