DOCSLIB.ORG

Topoisomerase II

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Topoisomerase II Topoisomerase II-␣ Expression in Different Cell Cycle Phases in Fresh Human Breast Carcinomas Kenneth Villman, M.D., Elisabeth Ståhl, M.S., Göran Liljegren, M.D., Ph.D., Ulf Tidefelt, M.D., Ph.D., Mats G. Karlsson, M.D., Ph.D. Departments of Oncology (KV), Pathology (ES, MGK), Surgery (GL), and Medicine (UT), Örebro University Hospital, Örebro, Sweden; and Karolinska Institute (UT), Stockholm, Sweden cluded in most adjuvant chemotherapy regimens Topoisomerase II-␣ (topo II␣) is the key target en- for breast cancer. Anthracyclines belong to the an- zyme for the topoisomerase inhibitor class of anti- ticancer agents called topoisomerase (topo) II cancer drugs. In normal cells, topo II␣ is expressed inhibitors. predominantly in the S/G2/M phase of the cell cycle. Topoisomerases are enzymes, present in the nu- In malignant cells, in vitro studies have indicated clei in mammalian cells, that regulate topological that the expression of topo II␣ is both higher and changes in DNA that are vital for many cellular less dependent on proliferation state in the cell. We processes such as replication and transcription (5). studied fresh specimens from 50 cases of primary They perform their function by introducing tran- breast cancer. The expression of topo II␣ in differ- sient protein-bridged DNA breaks on one (topo- ent cell cycle phases was analyzed with two- isomerase I) or both DNA strands (topoisomerase parameter flow cytometry using the monoclonal II; 6). There are two isoenzymes of topoisomerase antibody SWT3D1 and propidium iodide staining. II, with genetically and biochemical distinct fea- ␣ The expression of topo II was significantly higher tures. Topoisomerase II-␣ (topo II␣), with a molec- in the S/G2/M phase of the cell cycle than in the ular weight of 170 kd, is located on chromosome 17 G0/G1 phase in both DNA diploid and DNA nondip- (7), and topoisomerase II-␤ (topo II␤), with a mo- loid tumors. In 18 of 21 diploid tumors, and in 25 of lecular weight of 180 kd, is located on chromosome ␣ 29 nondiploid tumors, >50% of the topo II –posi- 3 (8). Topo II␣ is the primary drug target for anthra- tive cells were in the G0/G1 phase. This significant cyclines (6, 9). ␣ expression of topo II in the G0/G1 phase of the cell Topoisomerase II inhibitors inhibit the rejoining cycle may have clinically important implications for step in the breakage-rejoining cycle of topo II␣ and treatment efficacy of topoisomerase II inhibitors. thereby shift the equilibrium toward a key covalent reaction intermediate termed the cleavable complex KEY WORDS: Breast cancer, Cell cycle, DNA flow (10). This leads to double-stranded DNA breaks, ␣ cytometry, Topoisomerase II . which can, by a partly unknown mechanism, lead to Mod Pathol 2002;15(5):486–491 cell death (11). In vitro studies on cell lines from different human malignancies has shown that sensi- Breast cancer is the most common cancer among tivity to topoisomerase II inhibitors is dependent on women in Sweden and in the United States, and the the cellular level of topo II␣ (12–14). disease is fatal to every fourth woman despite wide Topo II␤ is expressed relatively constantly through- use of adjuvant chemotherapy (1–3). The limited out the cell cycle in both normal and transformed efficacy of chemotherapy is due to intrinsic or ac- cells (15, 16). In cell lines, it has been shown that in quired resistance to the drugs in use. Anthracy- normal cells, topo II␣ is a marker of proliferation and clines are the most active chemotherapeutic agents that expression is restricted to S and G2/M phases of in advanced breast cancer (4) and are today in- the cell cycle, whereas in transformed cells, the topo II␣ expression is both higher and less dependent on proliferation state in the cell (15, 17–21). The altered Copyright © 2002 by The United States and Canadian Academy of topo II␣ distribution may be caused by prolonged Pathology, Inc. VOL. 15, NO. 5, P. 486, 2002 Printed in the U.S.A. intracellular half-life of topo II␣ associated with the Date of acceptance: January 23, 2002. This study was supported by grants from the Research Committee of malignant transformation (17). This dysregulation of Örebro County Council, Sweden. the expression of topo II␣ in the cell cycle in malig- Address reprint requests to: Kenneth Villman, M.D., Department of On- cology, Örebro University Hospital, S-701 85 Örebro, Sweden; e-mail: nant cells may have implications for the efficacy of [email protected]; fax: 46-19-101768. treatment with topoisomerase II inhibitors. 486 There is today little information regarding topo pared within 18 hours of excision. Fresh breast tis- II␣ expression related to different cell cycle phases sue was mechanically disaggregated with Medima- in human neoplasms. A study on ovarian cancer chine (ConsulT.S., Rivalta di Torino, Italy) in 0.05 M with two-parameter flow cytometric analysis has PBS for 1–2 minutes. The cell suspension was fil- detected topo II␣–positive cells in the G1 phase in tered through a 40-␮m filter and centrifuged at 300 14 of 29 cases (22). ϫ g for 5 minutes. After one wash in PBS the cells The aim of the present study was to examine the were diluted to a concentration of 10 ϫ 106/mL in distribution of topo II␣ in the different cell cycle 0.05 M PBS. phases with two-parameter flow cytometry (FCM) in fresh human breast carcinomas. Determination of S-Phase Fraction and Ploidy A regular DNA staining with DNAcon 3 tubes MATERIALS AND METHODS (ConsulT.S.) was performed. The tubes contained a dehydrated buffer (lysing agent, RNAse and croma- Patients tin stabiliser). Propidium iodide solution (PI) was Material from 50 tumors from 49 women oper- added and the tube left at room temperature for 5 ated on at Örebro University Hospital was used in minutes in the dark. Two different control cells, this study. All breast cancer samples where sent chicken and salmon, were added to 1 ϫ 106 sample fresh from the operation theater to the Department cells and then incubated for 1 hour in an ice bath in of Pathology. Only patients with tumor size big the dark. enough to allow adequate material for routine his- The stained nuclei were analyzed in a FACScan tology, DNA analysis, estrogen receptor and proges- equipped with a 488-nm argon laser (Becton Dick- terone receptor determinations, and FCM for topo inson, Immunocytometry Systems, CA) with II␣ were included. CellQuest (Becton Dickinson) software. The DNA Patient age at operation ranged from 33 to 94 analysis was based on 15 000 events. To calculate years, with a median age of 61 years. All tumors DNA ploidy and S-phase fraction (SPF), Modfit were invasive breast carcinomas. Tumor character- (Becton Dickinson) software was used. Tumors istics are shown in Table 1. No patient had received were defined as either diploid (one stemline) or preoperative chemotherapy, hormonal therapy, or nondiploid (two or more cell populations; 23). The radiotherapy. Patients were given oral and written SPF was compared with that of the total number of information regarding the study, and informed cells expressing topo II␣. SPF was technically feasi- consent was obtained. ble in 82% (41/50) of the tumors. Preparation of Fresh Breast Tumor Tissue Flow Cytometry TopoII␣ Analysis The breast tumor samples were stored at 4° Cin Simultaneous measurement of topo II␣ and DNA ϫ 0.05 M phosphate buffered saline (PBS) and pre- content was done by two-parameter FCM. After 1 106 cells was added to each of two tubes, the tubes TABLE 1. Characteristics of the 50 Breast Carcinomas were incubated with 600-␮L of lysing solution (0.5% Number % Triton X-100 [Sigma Chemical Co, St. Louis, MO], Tumor size 1% BSA [Life Technologies, Paisley, Scotland], and pT1 16 32 0.2 ␮g/mL EDTA [Sigma] in 0.05 M PBS), pH 8.6. The pT2 32 64 cells were incubated for 10 minutes in an ice bath in pT3 2 4 Ϫ Histologic type the dark. The nuclei were fixed in 2 mL of 20° C Ductal 43 86 99.8% methanol pa (J.T. Baker. Deventer, Holland) Lobular 3 6 for 10 minutes in an ice-bath in the dark. After one Other 4 8 M Tumor gradea wash with 0.05 PBS, the tubes were centrifuged 1918for 5 minutes at 300 ϫ g. Topo II␣ clone SWT3D1 22550IgG1␬ (DAKO, Glostrup, Denmark; 1:100) was 31632 Estrogen receptor status added to one of the tubes. After incubation, a FITC- Positive 39 78 conjugated rat anti-mouse (RAM) antibody (DAKO; Negative 11 22 1:50) was added to both tubes and incubated, fol- Ploidy status ␮ Diploid 21 42 lowed by another incubation with 400 LofPI(10 Nondiploid 29 58 ␮g/mL; Sigma), with RNAse (200 ␮g/mL; Sigma). Axillary nodal status The last three incubations were performed for 30 pN0 26 52 pN1 22 44 minutes in an ice bath in the dark. NDb 24The stained nuclei were analyzed in a FACScan a Using the Elston modification of the Bloom and Richardson system. equipped with a 488-nm argon laser (Becton Dick- b ND, axillary node dissection not done. inson) and CellQuest (Becton Dickinson) software. TopoII␣ in Breast Cancer (K. Villman et al.) 487 FIGURE 1. Flow cytometric analysis of topo II␣ and DNA contents in fresh breast carcinomas. The DNA content of the investigated nuclei, reflecting the cell cycle phase, was determined after incubation with propidium iodide and RNAse. (A and D) show DNA histograms used for separation of the different cell cycle phases. A–C, tumor with diploid DNA content showing cells in G0/G1 phase (*) and S/G2/M phase (**).
Load more

Recommended publications
  • Selective DNA-Pkcs Inhibition Extends the Therapeutic Index of Localized Radiotherapy and Chemotherapy
    Selective DNA-PKcs inhibition extends the therapeutic index of localized radiotherapy and chemotherapy Catherine E. Willoughby, … , Anderson J. Ryan, Stephen R. Wedge J Clin Invest. 2019. https://doi.org/10.1172/JCI127483. Research Article Oncology Therapeutics Graphical abstract Find the latest version: https://jci.me/127483/pdf The Journal of Clinical Investigation RESEARCH ARTICLE Selective DNA-PKcs inhibition extends the therapeutic index of localized radiotherapy and chemotherapy Catherine E. Willoughby,1 Yanyan Jiang,2 Huw D. Thomas,1 Elaine Willmore,1 Suzanne Kyle,1 Anita Wittner,1 Nicole Phillips,1 Yan Zhao,1 Susan J. Tudhope,1 Lisa Prendergast,1 Gesa Junge,1 Luiza Madia Lourenco,2 M. Raymond V. Finlay,3 Paul Turner,3 Joanne M. Munck,4 Roger J. Griffin,5 Tommy Rennison,5 James Pickles,5 Celine Cano,5 David R. Newell,1 Helen L. Reeves,1,6 Anderson J. Ryan,2 and Stephen R. Wedge1 1Cancer Research UK Newcastle Drug Discovery Unit, Translational and Clinical Research Institute, Newcastle University Centre for Cancer, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, United Kingdom. 2Cancer Research UK and UK Medical Research Council Oxford Institute for Radiation Oncology, Department of Oncology, University of Oxford, Oxford, United Kingdom. 3Medicinal Chemistry, Oncology, IMED Biotech Unit, AstraZeneca, Cambridge, United Kingdom. 4Astex Pharmaceuticals, Cambridge, United Kingdom. 5Cancer Research UK Newcastle Drug Discovery Unit, Chemistry, School of Natural and Environmental Sciences, Newcastle University, Newcastle upon Tyne, United Kingdom. 6Hepatopancreatobiliary Multidisciplinary Team, Freeman Hospital, Newcastle upon Tyne Hospitals NHS Foundation Trust, Newcastle upon Tyne, United Kingdom. Potentiating radiotherapy and chemotherapy by inhibiting DNA damage repair is proposed as a therapeutic strategy to improve outcomes for patients with solid tumors.
    [Show full text]
  • Up-Regulation of Telomerase Activity in Human Pancreatic Cancer Cells After Exposure to Etoposide
    British Journal of Cancer (2000) 82(11), 1819–1826 © 2000 Cancer Research Campaign DOI: 10.1054/ bjoc.2000.1117, available online at http://www.idealibrary.com on Up-regulation of telomerase activity in human pancreatic cancer cells after exposure to etoposide N Sato, K Mizumoto, M Kusumoto, S Nishio, N Maehara, T Urashima, T Ogawa and M Tanaka Department of Surgery and Oncology, Graduate School of Medical Sciences, Kyushu University, Fukuoka 812-8582, Japan Summary Telomerase plays a critical role in the development of cellular immortality and oncogenesis. Activation of telomerase occurs in a majority of human malignant tumours, and the relation between telomerase and vulnerability to drug-mediated apoptosis remains unclear. In this study, we demonstrate, for the first time, up-regulation of telomerase activity in human pancreatic cancer cells treated with etoposide, a topoisomerase II inhibitor. Exposure of MIA PaCa-2 cells to etoposide at various concentrations (1–30 µM) resulted in two- to threefold increases in telomerase activity. Up-regulation was detectable 24 h after drug exposure and was accompanied by enhanced expression of mRNA of the human telomerase reverse transcriptase. Telomerase activation was also observed in AsPC-1 and PANC-1 cells but not in KP-3 and KP-1N cells. Furthermore, we found a negative correlation between increased telomerase activity and the percentage of dead cells after etoposide treatment. These findings suggest the existence of an anti-apoptotic pathway through which telomerase is up-regulated in response to DNA damage. This telomerase activation pathway may be one of the mechanisms responsible for the development of etoposide resistance in certain pancreatic cancer cells.
    [Show full text]
  • Anti-Neoplastic Effects of Topoisomerase Inhibitors in Canine Mammary Carcinoma, Melanoma, and Osteosarcoma Cell Title Lines
    Anti-neoplastic effects of topoisomerase inhibitors in canine mammary carcinoma, melanoma, and osteosarcoma cell Title lines Ong, Siew Mei; Yamamoto, Hiroki; Saeki, Kohei; Tanaka, Yuiko; Yoshitake, Ryohei; Nishimura, Ryohei; Nakagawa, Author(s) Takayuki Citation Japanese Journal of Veterinary Research, 65(1), 17-28 Issue Date 2017-02 DOI 10.14943/jjvr.65.1.17 Doc URL http://hdl.handle.net/2115/64788 Type bulletin (article) File Information 65-1_017-028.pdf Instructions for use Hokkaido University Collection of Scholarly and Academic Papers : HUSCAP Japanese Journal of Veterinary Research 65(1): 17-28, 2017 REGULAR PAPER Experimental Research Anti-neoplastic effects of topoisomerase inhibitors in canine mammary carcinoma, melanoma, and osteosarcoma cell lines Siew Mei Ong1,†), Hiroki Yamamoto1,†), Kohei Saeki1), Yuiko Tanaka1), Ryohei Yoshitake1), Ryohei Nishimura1) and Takayuki Nakagawa1,＊) 1) Laboratory of Veterinary Surgery, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1, Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan Received for publication, November 29, 2016; accepted, December 27, 2016 Abstract Numerous topoisomerase inhibitors with proven efficacy have been used extensively to treat various human neoplasms. However, among these, only doxorubicin has been used and studied extensively in veterinary oncology. The current study was performed to evaluate the responsiveness of canine osteosarcoma (cOSA), mammary gland tumour (cMGT), and malignant melanoma (cMM) cell lines to several topoisomerase inhibitors. In addition, the correlation between the sensitivity to treatment and multi-drug resistant (MDR) factors was investigated. cOSA cell lines exhibited higher sensitivity than cMGT and cMM cell lines to all the topoisomerase inhibitors tested in vitro; this was associated with the levels of multi-drug resistance protein 1 (MDR1) gene expression in the cOSA cell lines.
    [Show full text]
  • CLINICAL and PHARMACOLOGICAL STUDIES on the TOPOISOMERASE I INHIBITOR TOPOTECAN Cover Design: G.L.M
    CLINICAL AND PHARMACOLOGICAL STUDIES ON THE TOPOISOMERASE I INHIBITOR TOPOTECAN Cover design: G.L.M. Obbens Lay-out: A.A.C. Treuren-den Braber Printed by: Copynomie Erasmus Multicopy Facility Management B.V. Burg. Oudlaan 50 3062 PA Rotterdam The Netherlands phone: 010-4082898 fax: 010-4528183 ISBN: 9090091718 Copyright: G.J. Creemers. All rights reserved. No part of this publication may be reproduced, stored in a retrieval system, or transmitted in any form or by any means, mechanically, by photocopying, by recording or otherwise without the prior permission of the author. Clinical and pharmacological studies on the topoisomerase I inhibitor topotecan Klinisch en farmacologisch onderzoek met de topoisomerase I rammer topotecan. PROEFSCHRIFT ter verkrijging van de graad van doctor aan de Erasmus Universiteit Rotterdam op gezag van de rector magnificus Prof. dr. P.W.C. Akkermans M.A. en volgens besluit van het college voor promoties. De open bare verdediging zal plaatsvinden op woensdag 13 maart 1996 om 13.45 uur door Gerardus Johannes Marie Creemers geboren te Eindhoven PROMOTIECOMMISSIE Promotor: Prof. dr. G. Stoter Copromotor: Dr. J. Verweij Overige leden: Prof. dr. D.O. Von Hoff Prof. dr. J.H. Beijnen Prof. dr. J.W. Oosterhuis Prof. dr. A. Hagenbeek "Aileen wat niet ophoudt pijn te doen, blijft in de herinnering" Contents Chapter 1 Introduction to the thesis. 1 Chapter 2 Review: The topoisomerase I inhibitors topotecan and irinotecan. 5 Cancer Treatment Reviews 1994: 20; 73·96. Chapter 3 Topotecan in colorectal cancer: A phase II study of the EORTe clinical trials group. 47 Annals of oncology 1995: 6; 844·846.
    [Show full text]
  • Differential Effects of the Poly (ADP-Ribose)Polymerase (PARP
    British Journal of Cancer (2001) 84(1), 106–112 © 2001 Cancer Research Campaign doi: 10.1054/ bjoc.2000.1555, available online at http://www.idealibrary.com on http://www.bjcancer.com Differential effects of the poly (ADP-ribose) polymerase (PARP) inhibitor NU1025 on topoisomerase I and II inhibitor cytotoxicity in L1210 cells in vitro KJ Bowman*, DR Newell, AH Calvert and NJ Curtin Cancer Research Unit, University of Newcastle upon Tyne Medical School, Framlington Place, Newcastle upon Tyne NE2 4HH, UK Summary The potent novel poly(ADP-ribose) polymerase (PARP) inhibitor, NU1025, enhances the cytotoxicity of DNA-methylating agents and ionizing radiation by inhibiting DNA repair. We report here an investigation of the role of PARP in the cellular responses to inhibitors of topoisomerase I and II using NU1025. The cytotoxicity of the topoisomerase I inhibitor, camptothecin, was increased 2.6-fold in L1210 cells by co-incubation with NU1025. Camptothecin-induced DNA strand breaks were also increased 2.5-fold by NU1025 and exposure to camptothecin-activated PARP. In contrast, NU1025 did not increase the DNA strand breakage or cytotoxicity caused by the topoisomerase II inhibitor etoposide. Exposure to etoposide did not activate PARP even at concentrations that caused significant levels of apoptosis. Taken together, these data suggest that potentiation of camptothecin cytotoxicity by NU1025 is a direct result of increased DNA strand breakage, and that activation of PARP by camptothecin-induced DNA damage contributes to its repair and consequently cell survival. However, in L1210 cells at least, it would appear that PARP is not involved in the cellular response to etoposide-mediated DNA damage.
    [Show full text]
  • Functional Genomics Approaches to Elucidate Vulnerabilities of Intrinsic and Acquired Chemotherapy Resistance
    cells Review Functional Genomics Approaches to Elucidate Vulnerabilities of Intrinsic and Acquired Chemotherapy Resistance Ronay Cetin 1,† , Eva Quandt 2,† and Manuel Kaulich 1,3,4,* 1 Institute of Biochemistry II, Goethe University Frankfurt-Medical Faculty, University Hospital, 60590 Frankfurt am Main, Germany; [email protected] 2 Faculty of Medicine and Health Sciences, Universitat Internacional de Catalunya, 08195 Barcelona, Spain; [email protected] 3 Frankfurt Cancer Institute, 60596 Frankfurt am Main, Germany 4 Cardio-Pulmonary Institute, 60590 Frankfurt am Main, Germany * Correspondence: [email protected]; Tel.: +49-(0)-69-6301-5450 † These authors contributed equally to this work. Abstract: Drug resistance is a commonly unavoidable consequence of cancer treatment that results in therapy failure and disease relapse. Intrinsic (pre-existing) or acquired resistance mechanisms can be drug-specific or be applicable to multiple drugs, resulting in multidrug resistance. The presence of drug resistance is, however, tightly coupled to changes in cellular homeostasis, which can lead to resistance-coupled vulnerabilities. Unbiased gene perturbations through RNAi and CRISPR technologies are invaluable tools to establish genotype-to-phenotype relationships at the genome scale. Moreover, their application to cancer cell lines can uncover new vulnerabilities that are associated with resistance mechanisms. Here, we discuss targeted and unbiased RNAi and CRISPR efforts in the discovery of drug resistance mechanisms by focusing on first-in-line chemotherapy and their enforced vulnerabilities, and we present a view forward on which measures should be taken to accelerate their clinical translation. Citation: Cetin, R.; Quandt, E.; Kaulich, M. Functional Genomics Keywords: chemotherapy resistance; cancer and drug vulnerabilities; functional genomics; RNAi Approaches to Elucidate Vulnerabilities of Intrinsic and and CRISPR screens Acquired Chemotherapy Resistance.
    [Show full text]
  • As New Horizons in Antifungal Treatment
    molecules Review ‘Acridines’ as New Horizons in Antifungal Treatment Iwona Gabriel Department of Pharmaceutical Technology and Biochemistry, Gda´nskUniversity of Technology, 80-233 Gda´nsk, Poland; [email protected]; Tel.: +48-58-3486-078; Fax: +48-58-3471-144 Received: 18 February 2020; Accepted: 23 March 2020; Published: 25 March 2020 Abstract: Frequent fungal infections in immunocompromised patients and mortality due to invasive mycosis are important clinical problems. Opportunistic pathogenic Candida species remain one of the leading causes of systemic mycosis worldwide. The repertoire of antifungal chemotherapeutic agents is very limited. Although new antifungal drugs such as lanosterol 14α-demethylase and β-glucan synthase inhibitors have been introduced into clinical practice, the development of multidrug resistance has become increasingly significant. The urgency to expand the range of therapeutic options for the treatment of fungal infections has led researchers in recent decades to seek alternative antifungal targets to the conventional ones currently used. Among them, many compounds containing an acridine scaffold have been synthesized and tested. In this review, the applicability of acridines and their functional analogues acridones as antifungal agents is described. Acridine derivatives usage in photoantifungal chemotherapy, interactions with fungal transporters resulting in modulation of efflux/influx pumps and the effect of acridine derivatives on fungal topoisomerases are discussed. This article explores new perspectives
    [Show full text]
  • Potentiation of Topoisomerase Inhibitor-Induced DNA Strand Breakage and Cytotoxicity by Tumor Necrosis Factor: Enhancement of To
    [CANCER RESEARCH 50. 2636-2640, May I. 1990] Potentiation of Topoisomerase Inhibitor-induced DNA Strand Breakage and Cytotoxicity by Tumor Necrosis Factor: Enhancement of Topoisomerase Activity as a Mechanism of Potentiation Teruhiro Utsugi, Michael R. Mattern, Christopher K. Mirabelli, and Nabil Hanna1 Departments of Immunology [T. I'., ,V. H.¡and Molecular Pharmacology [M. R. M., C. K. M.¡,Smith Kline & French Laboratories, King of Prussia, Pennsylvania 19406- 0939 ABSTRACT onstrated that treatment of target cells with topoisomerase I or II inhibitors enhanced their killing by natural cytotoxic cell- A combination of tumor necrosis factor (INI) and the topoisomerase mediated cytotoxicity (8). I inhibitor, camptothecin, or the topoisomerase II inhibitors, teniposide and amsacrine, produced dose-dependent synergistic cytotoxicity against Topoisomerases are nuclear enzymes which catalyze the for the murine L929 fibrosarcoma cells. Similar synergy was not observed mation of various topological isomers of DNA by transiently breaking and rejoining DNA (9-12). Topoisomerase I catalyzes with a combination of TNF and bleomycin. To define the role of TNF in the augmentation of tumor cell killing by topoisomerase I or II inhibitors, single strand breakage and strand passage independent of ATP, the effect of TNF on the production of enzyme-linked DNA strand breaks whereas topoisomerase II catalyzes double strand DNA break induced in cells by topoisomerase inhibitors was investigated. L929 cells age and strand passage in the presence of ATP. Topoisomerase incubated for l h with the topoisomerase inhibitors contained protein- inhibitors are believed to induce cytotoxicity by stabilizing linked strand breaks. In contrast, INI alone did not induce DNA strand covalent complexes of enzyme and strand-cleaved DNA, which breakage.
    [Show full text]
  • DNA-PK Inhibition by NU7441 Enhances Chemosensitivity to Topoisomerase Inhibitor in Non-Small Cell Lung Carcinoma Cells by Blocking DNA Damage Repair
    Yonago Acta Medica 2017;60:09–15 Original Article DNA-PK Inhibition by NU7441 Enhances Chemosensitivity to Topoisomerase Inhibitor in Non-Small Cell Lung Carcinoma Cells by Blocking DNA Damage Repair Masaaki Yanai, Haruhiko Makino, Bingqiong Ping, Kenichi Takeda, Natsumi Tanaka, Tomohiro Sakamoto, Kosuke Yamaguchi, Masahiro Kodani, Akira Yamasaki, Tadashi Igishi and Eiji Shimizu Division of Medical Oncology and Molecular Respirology, Department of Multidisciplinary Internal Medicine, School of Medicine, Tot- tori University Faculty of Medicine, Yonago 683-8504, Japan ABSTRACT cause of cancer-related death worldwide. Over half of Background DNA double-strand breaks (DSBs) are patients with NSCLC have been advanced or metastatic the most cytotoxic form of DNA damage and are in- lesions at the time of diagnosis.1 Systemic chemotherapy duced by ionizing radiation and specific chemotherapeu- is the standard treatment for patients with advanced NS- tic agents, such as topoisomerase inhibitors. Cancer cells CLC. In many countries, including Japan, cytotoxic che- acquire resistance to such therapies by repairing DNA motherapeutic agents are used as the first-line treatment DSBs. A major pathway for the repair of DNA DSBs is for advanced NSCLC patients. However, these cytotoxic non-homologous end-joining (NHEJ), which requires chemotherapies have reached a therapeutic plateau and DNA-dependent protein kinase (DNA-PK) activity. In have limited survival benefits for advanced NSCLC pa- this study, we investigated the effect of NU7441, a syn- tients. In recent years, treatment with targeted agents has thetic small-molecule compound, as a specific inhibitor markedly improved progression-free and overall survival of DNA-PK on the chemosensitization of non-small cell in patients with advanced NSCLC harboring EGFR mu- lung carcinoma (NSCLC) A549 cells.
    [Show full text]
  • RECQ5-Dependent Sumoylation of DNA Topoisomerase I Prevents Transcription-Associated Genome Instability
    ARTICLE Received 20 Aug 2014 | Accepted 23 Feb 2015 | Published 8 Apr 2015 DOI: 10.1038/ncomms7720 RECQ5-dependent SUMOylation of DNA topoisomerase I prevents transcription-associated genome instability Min Li1, Subhash Pokharel1,*, Jiin-Tarng Wang1,*, Xiaohua Xu1,* & Yilun Liu1 DNA topoisomerase I (TOP1) has an important role in maintaining DNA topology by relaxing supercoiled DNA. Here we show that the K391 and K436 residues of TOP1 are SUMOylated by the PIAS1–SRSF1 E3 ligase complex in the chromatin fraction containing active RNA polymerase II (RNAPIIo). This modification is necessary for the binding of TOP1 to RNAPIIo and for the recruitment of RNA splicing factors to the actively transcribed chromatin, thereby reducing the formation of R-loops that lead to genome instability. RECQ5 helicase promotes TOP1 SUMOylation by facilitating the interaction between PIAS1, SRSF1 and TOP1. Unexpectedly, the topoisomerase activity is compromised by K391/K436 SUMOylation, and this provides the first in vivo evidence that TOP1 activity is negatively regulated at transcriptionally active chromatin to prevent TOP1-induced DNA damage. Therefore, our data provide mechanistic insight into how TOP1 SUMOylation contributes to genome maintenance during transcription. 1 Department of Radiation Biology, Beckman Research Institute, City of Hope, Duarte, California 91010-3000, USA. * These authors contributed equally to this work. Correspondence and requests for materials should be addressed to Y.L. (email: [email protected]). NATURE COMMUNICATIONS | 6:6720 | DOI: 10.1038/ncomms7720 | www.nature.com/naturecommunications 1 & 2015 Macmillan Publishers Limited. All rights reserved. ARTICLE NATURE COMMUNICATIONS | DOI: 10.1038/ncomms7720 he prevention and efficient repair of DNA double-stranded transcriptionally active chromatin to prevent R-loops.
    [Show full text]
  • In Vitro Synergistic Effects of Anthracycline Antitumor Agents And
    evelo f D pin l o g a D Matsumoto et al., J Develop Drugs 2014, 3:2 n r r u u g o s J Journal of Developing Drugs DOI: 10.4172/2329-6631.1000125 ISSN: 2329-6631 Research Article Open Access In vitro Synergistic Effects of Anthracycline Antitumor Agents and Fluconazole Against Azole-Resistant Candida albicans Clinical Isolates Matsumoto S, Kurakado S, Shiokama T, Ando Y, Aoki N, Cho O and Sugita T* Department of Microbiology, Meiji Pharmaceutical University, Kiyose, Tokyo Japan Abstract The number of azole-resistant Candida albicans clinical isolates is increasing. This study searched for compounds that are functionally synergistic with fluconazole against azole-resistant C. albicans strains. Synergistic effects were evaluated using the checkerboard method in a time-kill study using anthracycline antitumor agents and azole-resistant C. albicans strains. Of the five anthracycline agents examined, aclarubicin by itself had antifungal effects, whereas daunorubicin, doxorubicin, epirubicin, and idarubicin did not show antifungal effects alone, but did exert dose- and time- dependent synergistic effects with fluconazole against the C. albicans strains. No antitumor agent other than anthracycline exhibited an anti-Candida effect. Anthracycline compounds may therefore be useful as seeds for development of new antifungal agents. Keywords: Anthracycline; antitumor agent; azole-resistant Candida Materials and Methods albicans; synergy Strains used Introduction Twelve azole-resistant Candida albicans strains obtained from Candida albicans is the most frequently observed opportunistic patients’ blood were examined in this study. All strains were cultured fungal pathogen and causes deep-seated fungal infection, mainly on Sabouraud dextrose agar (SDA) plates at 37°C.
    [Show full text]
  • Type II DNA Topoisomerases Cause Spontaneous Double-Strand Breaks in Genomic DNA
    G C A T T A C G G C A T genes Review Type II DNA Topoisomerases Cause Spontaneous Double-Strand Breaks in Genomic DNA Suguru Morimoto 1, Masataka Tsuda 1, Heeyoun Bunch 2 , Hiroyuki Sasanuma 1, Caroline Austin 3 and Shunichi Takeda 1,* 1 Department of Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshida Konoe, Sakyo-ku, Kyoto 606-8501, Japan; [email protected] (S.M.); [email protected] (M.T.); [email protected] (H.S.) 2 Department of Applied Biosciences, College of Agriculture and Life Sciences, Kyungpook National University, Daegu 41566, Korea; [email protected] 3 The Institute for Cell and Molecular Biosciences, the Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne NE2 4HH, UK; [email protected] * Correspondence: [email protected]; Tel.: +81-(075)-753-4411 Received: 30 September 2019; Accepted: 26 October 2019; Published: 30 October 2019 Abstract: Type II DNA topoisomerase enzymes (TOP2) catalyze topological changes by strand passage reactions. They involve passing one intact double stranded DNA duplex through a transient enzyme-bridged break in another (gated helix) followed by ligation of the break by TOP2. A TOP2 poison, etoposide blocks TOP2 catalysis at the ligation step of the enzyme-bridged break, increasing the number of stable TOP2 cleavage complexes (TOP2ccs). Remarkably, such pathological TOP2ccs are formed during the normal cell cycle as well as in postmitotic cells. Thus, this ‘abortive catalysis’ can be a major source of spontaneously arising DNA double-strand breaks (DSBs).
    [Show full text]