Additive effects of a small molecular PCNA inhibitor PCNA-I1S and DNA damaging agents on growth inhibition and DNA damage in prostate and lung cancer cells


Autoři: Shan Lu aff001;  Zhongyun Dong aff001
Působiště autorů: Division of Hematology-Oncology, Department of Internal Medicine, University of Cincinnati College of Medicine, Cincinnati, Ohio, United States of America aff001
Vyšlo v časopise: PLoS ONE 14(10)
Kategorie: Research Article
doi: 10.1371/journal.pone.0223894

Souhrn

Proliferating cell nuclear antigen (PCNA) is essential for DNA replication and repair, and cell growth and survival. Previously, we identified a novel class of small molecules that bind directly to PCNA, stabilize PCNA trimer structure, reduce chromatin-associated PCNA, selectively inhibit tumor cell growth, and induce apoptosis. The purpose of this study was to investigate the combinatorial effects of lead compound PCNA-I1S with DNA damaging agents on cell growth, DNA damage, and DNA repair in four lines of human prostate and lung cancer cells. The DNA damage agents used in the study include ionizing radiation source cesium-137 (Cs-137), chemotherapy drug cisplatin (cisPt), ultraviolet-C (UV-C), and oxidative compound H2O2. DNA damage was assessed using immunofluorescent staining of γH2AX and the Comet assay. The homologous recombination repair (HRR) was determined using a plasmid-based HRR reporter assay and the nucleotide excision repair (NER) was indirectly examined by the removal of UV-induced cyclobutane pyrimidine dimers (CPD). We found that PCNA-I1S inhibited cell growth in a dose-dependent manner and significantly enhanced the cell growth inhibition induced by pretreatment with DNA damaging agents Cs-137 irradiation, UV-C, and cisPt. However, the additive growth inhibitory effects were not observed in cells pre-treated with PCNA-I1S, followed by treatment with cisPt. H2O2 enhanced the level of chromatin-bound PCNA in quiescent cells, which was attenuated by PCNA-I1S. DNA damage was induced in cells treated with either PCNA-I1S or cisPt alone and was significantly elevated in cells exposed to the combination of PCNA-I1S and cisPt. Finally, PCNA-I1S attenuated repair of DNA double strand breaks (DSBs) by HRR and the removal of CPD by NER. These data suggest that targeting PCNA with PCNA-I1S may provide a novel approach for enhancing the efficacy of chemotherapy and radiation therapy in treatment of human prostate and lung cancer.

Klíčová slova:

Cell growth – Cell staining – DAPI staining – DNA damage – DNA repair – DNA replication – Immunofluorescence staining – Ultraviolet C


Zdroje

1. Almendral JM, Huebsch D, Blundell PA, Macdonald-Bravo H, Bravo R. Cloning and sequence of the human nuclear protein cyclin: homology with DNA-binding proteins. Proc Natl Acad Sci U S A. 1987;84(6):1575–9. doi: 10.1073/pnas.84.6.1575 2882507.

2. Stoimenov I, Helleday T. PCNA on the crossroad of cancer. Biochemical Society transactions. 2009;37(Pt 3):605–13. doi: 10.1042/BST0370605 19442257.

3. Lu S, Lee J, Revelo M, Wang X, Lu S, Dong Z. Smad3 is overexpressed in advanced human prostate cancer and necessary for progressive growth of prostate cancer cells in nude mice. Clin Cancer Res. 2007;13(19):5692–702. doi: 10.1158/1078-0432.CCR-07-1078 17908958.

4. Harper ME, Glynne-Jones E, Goddard L, Wilson DW, Matenhelia SS, Conn IG, et al. Relationship of proliferating cell nuclear antigen (PCNA) in prostatic carcinomas to various clinical parameters. Prostate. 1992;20(3):243–53. doi: 10.1002/pros.2990200309 1374182.

5. Zdunek M, Korobowicz E. Expression of PCNA in non-small cell lung cancer before and after treatment with cisplatin and vepeside. Polish journal of pathology: official journal of the Polish Society of Pathologists. 2000;51(2):77–81. Epub 2000/09/07. 10974930.

6. Kimos MC, Wang S, Borkowski A, Yang GY, Yang CS, Perry K, et al. Esophagin and proliferating cell nuclear antigen (PCNA) are biomarkers of human esophageal neoplastic progression. Int J Cancer. 2004;111(3):415–7. doi: 10.1002/ijc.20267 15221970.

7. Cappello F, Ribbene A, Campanella C, Czarnecka AM, Anzalone R, Bucchieri F, et al. The value of immunohistochemical research on PCNA, p53 and heat shock proteins in prostate cancer management: a review. Eur J Histochem. 2006;50(1):25–34. 16584982.

8. Malkas LH, Herbert BS, Abdel-Aziz W, Dobrolecki LE, Liu Y, Agarwal B, et al. A cancer-associated PCNA expressed in breast cancer has implications as a potential biomarker. Proc Natl Acad Sci U S A. 2006;103(51):19472–7. doi: 10.1073/pnas.0604614103 17159154.

9. Helleday T, Petermann E, Lundin C, Hodgson B, Sharma RA. DNA repair pathways as targets for cancer therapy. Nature Reviews: Cancer. 2008;8(3):193–204. Epub 2008/02/08. doi: 10.1038/nrc2342 18256616.

10. Hosoya N, Miyagawa K. Targeting DNA damage response in cancer therapy. Cancer Sci. 2014;105(4):370–88. doi: 10.1111/cas.12366 24484288.

11. Naryzhny SN. Proliferating cell nuclear antigen: a proteomics view. Cell Mol Life Sci. 2008;65(23):3789–808. doi: 10.1007/s00018-008-8305-x 18726183.

12. Krishna TS, Kong XP, Gary S, Burgers PM, Kuriyan J. Crystal structure of the eukaryotic DNA polymerase processivity factor PCNA. Cell. 1994;79(7):1233–43. doi: 10.1016/0092-8674(94)90014-0 8001157.

13. Bowman GD, O'Donnell M, Kuriyan J. Structural analysis of a eukaryotic sliding DNA clamp-clamp loader complex. Nature. 2004;429(6993):724–30. doi: 10.1038/nature02585 15201901.

14. Fukuda K, Morioka H, Imajou S, Ikeda S, Ohtsuka E, Tsurimoto T. Structure-function relationship of the eukaryotic DNA replication factor, proliferating cell nuclear antigen. J Biol Chem. 1995;270(38):22527–34. Epub 1995/09/22. doi: 10.1074/jbc.270.38.22527 7673244.

15. Moldovan GL, Pfander B, Jentsch S. PCNA, the maestro of the replication fork. Cell. 2007;129(4):665–79. doi: 10.1016/j.cell.2007.05.003 17512402.

16. Maga G, Hubscher U. Proliferating cell nuclear antigen (PCNA): a dancer with many partners. J Cell Sci. 2003;116(Pt 15):3051–60. doi: 10.1242/jcs.00653 12829735

17. Witko-Sarsat V, Mocek J, Bouayad D, Tamassia N, Ribeil JA, Candalh C, et al. Proliferating cell nuclear antigen acts as a cytoplasmic platform controlling human neutrophil survival. J Exp Med. 2010;207(12):2631–45. doi: 10.1084/jem.20092241 20975039; PubMed Central PMCID: PMC2989777.

18. Ohayon D, De Chiara A, Chapuis N, Candalh C, Mocek J, Ribeil JA, et al. Cytoplasmic proliferating cell nuclear antigen connects glycolysis and cell survival in acute myeloid leukemia. Sci Rep. 2016;6:35561. Epub 2016/10/21. doi: 10.1038/srep35561 27759041; PubMed Central PMCID: PMC5069676.

19. Gilljam KM, Feyzi E, Aas PA, Sousa MM, Muller R, Vagbo CB, et al. Identification of a novel, widespread, and functionally important PCNA-binding motif. The Journal of cell biology. 2009;186(5):645–54. Epub 2009/09/09. doi: 10.1083/jcb.200903138 19736315; PubMed Central PMCID: PMC2742182.

20. Horton NC, Mathew SO, Mathew PA. Novel interaction between proliferating cell nuclear antigen and HLA I on the surface of tumor cells inhibits NK cell function through NKp44. PloS one. 2013;8(3):e59552. doi: 10.1371/journal.pone.0059552 23527218; PubMed Central PMCID: PMC3602199.

21. Muller R, Misund K, Holien T, Bachke S, Gilljam KM, Vatsveen TK, et al. Targeting proliferating cell nuclear antigen and its protein interactions induces apoptosis in multiple myeloma cells. PloS one. 2013;8(7):e70430. Epub 2013/08/13. doi: 10.1371/journal.pone.0070430 23936203; PubMed Central PMCID: PMC3729839.

22. Sogaard CK, Moestue SA, Rye MB, Kim J, Nepal A, Liabakk NB, et al. APIM-peptide targeting PCNA improves the efficacy of docetaxel treatment in the TRAMP mouse model of prostate cancer. Oncotarget. 2018;9(14):11752–66. doi: 10.18632/oncotarget.24357 29545934; PubMed Central PMCID: PMC5837745.

23. Smith SJ, Gu L, Phipps EA, Dobrolecki L, Mabrey KS, Gulley P, et al. A peptide mimicking a region in proliferating cell nuclear antigen specific to key protein interactions Is cytotoxic to breast cancer. Mol Pharmacol. 2015;87:263–76. doi: 10.1124/mol.114.093211 25480843

24. Actis M, Inoue A, Evison B, Perry S, Punchihewa C, Fujii N. Small molecule inhibitors of PCNA/PIP-box interaction suppress translesion DNA synthesis. Bioorganic & medicinal chemistry. 2013;21(7):1972–7. Epub 2013/02/12. doi: 10.1016/j.bmc.2013.01.022 23395113.

25. Punchihewa C, Inoue A, Hishiki A, Fujikawa Y, Connelly M, Evison B, et al. Identification of a small molecule PCNA inhibitor that disrupts interactions with PIP-Box proteins and inhibits DNA replication. Journal of Biological Chemistry. 2012;287(17):14289–300. Epub 2012/03/03. doi: 10.1074/jbc.M112.353201 22383522.

26. Gu L, Lingeman R, Yakushijin F, Sun E, Cui Q, Chao J, et al. The Anticancer Activity of a First-in-class Small-molecule Targeting PCNA. Clin Cancer Res. 2018;24(23):6053–65. Epub 2018/07/04. doi: 10.1158/1078-0432.CCR-18-0592 29967249; PubMed Central PMCID: PMC6279569.

27. Desplancq D, Freund G, Conic S, Sibler AP, Didier P, Stoessel A, et al. Targeting the replisome with transduced monoclonal antibodies triggers lethal DNA replication stress in cancer cells. Exp Cell Res. 2016;342(2):145–58. doi: 10.1016/j.yexcr.2016.03.003 26968636.

28. Gederaas OA, Sogaard CD, Viset T, Bachke S, Bruheim P, Arum CJ, et al. Increased Anticancer Efficacy of Intravesical Mitomycin C Therapy when Combined with a PCNA Targeting Peptide. Transl Oncol. 2014;7(6):812–23. doi: 10.1016/j.tranon.2014.10.005 25500092; PubMed Central PMCID: PMC4311026.

29. Tan Z, Wortman M, Dillehay KL, Seibel WL, Evelyn CR, Smith SJ, et al. Small-molecule targeting of proliferating cell nuclear antigen chromatin association inhibits tumor cell growth. Molecular pharmacology. 2012;81(6):811–9. Epub 2012/03/09. doi: 10.1124/mol.112.077735 22399488; PubMed Central PMCID: PMC3362894.

30. Dillehay KL, Seibel WL, Zhao D, Lu S, Dong Z. Target validation and structure-activity analysis of a series of novel PCNA inhibitors. Pharmacol Res Perspect. 2015;3(2):e00115. doi: 10.1002/prp2.115 25729582; PubMed Central PMCID: PMC4324689.

31. Dillehay K L., Lu S, Dong Z. Antitumor effects of a novel small molecule targeting PCNA chromatin association in prostate cancer. Mol Cancer Ther. 2014;13:2817–26. doi: 10.1158/1535-7163.MCT-14-0522 25253786

32. Tichy ED, Pillai R, Deng L, Liang L, Tischfield J, Schwemberger SJ, et al. Mouse embryonic stem cells, but not somatic cells, predominantly use homologous recombination to repair double-strand DNA breaks. Stem cells and development. 2010;19(11):1699–711. doi: 10.1089/scd.2010.0058 20446816; PubMed Central PMCID: PMC3128311.

33. Dong ZY, Ward NE, Fan D, Gupta KP, O'Brian CA. In vitro model for intrinsic drug resistance: effects of protein kinase C activators on the chemosensitivity of cultured human colon cancer cells. Mol Pharmacol. 1991;39(4):563–9. Epub 1991/04/01. 2017156.

34. Shibata Y, Nakamura T. Defective flap endonuclease 1 activity in mammalian cells is associated with impaired DNA repair and prolonged S phase delay. J Biol Chem. 2002;277(1):746–54. doi: 10.1074/jbc.M109461200 11687589.

35. Collins AR. The comet assay for DNA damage and repair: principles, applications, and limitations. Molecular biotechnology. 2004;26(3):249–61. doi: 10.1385/MB:26:3:249 15004294.

36. Gyori BM, Venkatachalam G, Thiagarajan PS, Hsu D, Clement MV. OpenComet: an automated tool for comet assay image analysis. Redox Biol. 2014;2:457–65. Epub 2014/03/14. doi: 10.1016/j.redox.2013.12.020 24624335; PubMed Central PMCID: PMC3949099.

37. Rastogi RP, Richa, Kumar A, Tyagi MB, Sinha RP. Molecular mechanisms of ultraviolet radiation-induced DNA damage and repair. J Nucleic Acids. 2010;2010:592980. Epub 2011/01/07. doi: 10.4061/2010/592980 21209706; PubMed Central PMCID: PMC3010660.

38. Soehnge H, Ouhtit A, Ananthaswamy ON. Mechanisms of induction of skin cancer by UV radiation. Front Biosci. 1997;2:d538–51. Epub 1997/10/31. doi: 10.2741/a211 9343491.

39. Reck M, Heigener DF, Mok T, Soria JC, Rabe KF. Management of non-small-cell lung cancer: recent developments. Lancet. 2013;382(9893):709–19. doi: 10.1016/S0140-6736(13)61502-0 23972814.

40. Strzalka W, Ziemienowicz A. Proliferating cell nuclear antigen (PCNA): a key factor in DNA replication and cell cycle regulation. Ann Bot. 2011;107(7):1127–40. Epub 2010/12/21. doi: 10.1093/aob/mcq243 21169293; PubMed Central PMCID: PMC3091797.

41. Karmakar P, Balajee AS, Natarajan AT. Analysis of repair and PCNA complex formation induced by ionizing radiation in human fibroblast cell lines. Mutagenesis. 2001;16(3):225–32. doi: 10.1093/mutage/16.3.225 11320148.

42. Savio M, Stivala LA, Bianchi L, Vannini V, Prosperi E. Involvement of the proliferating cell nuclear antigen (PCNA) in DNA repair induced by alkylating agents and oxidative damage in human fibroblasts. Carcinogenesis. 1998;19(4):591–6. doi: 10.1093/carcin/19.4.591 9600342.

43. Balajee AS, Dianova I, Bohr VA. Oxidative damage-induced PCNA complex formation is efficient in xeroderma pigmentosum group A but reduced in Cockayne syndrome group B cells. Nucleic Acids Res. 1999;27(22):4476–82. Epub 1999/10/28. doi: 10.1093/nar/27.22.4476 10536158; PubMed Central PMCID: PMC148732.

44. Blommaert FA, van Dijk-Knijnenburg HC, Dijt FJ, den Engelse L, Baan RA, Berends F, et al. Formation of DNA adducts by the anticancer drug carboplatin: different nucleotide sequence preferences in vitro and in cells. Biochemistry. 1995;34(26):8474–80. Epub 1995/07/04. doi: 10.1021/bi00026a031 7599137.

45. Kuo LJ, Yang LX. Gamma-H2AX—a novel biomarker for DNA double-strand breaks. In vivo (Athens, Greece). 2008;22(3):305–9. 18610740.

46. Rogakou EP, Pilch DR, Orr AH, Ivanova VS, Bonner WM. DNA double-stranded breaks induce histone H2AX phosphorylation on serine 139. Journal of Biological Chemistry. 1998;273(10):5858–68. Epub 1998/04/16. doi: 10.1074/jbc.273.10.5858 9488723.

47. Kinner A, Wu W, Staudt C, Iliakis G. Gamma-H2AX in recognition and signaling of DNA double-strand breaks in the context of chromatin. Nucleic acids research. 2008;36(17):5678–94. Epub 2008/09/06. doi: 10.1093/nar/gkn550 18772227; PubMed Central PMCID: PMC2553572.

48. Lee E, Oh E, Lee J, Sul D, Lee J. Use of the tail moment of the lymphocytes to evaluate DNA damage in human biomonitoring studies. Toxicol Sci. 2004;81(1):121–32. Epub 2004/06/05. doi: 10.1093/toxsci/kfh184 15178808.

49. Wit N, Buoninfante OA, van den Berk PC, Jansen JG, Hogenbirk MA, de Wind N, et al. Roles of PCNA ubiquitination and TLS polymerases kappa and eta in the bypass of methyl methanesulfonate-induced DNA damage. Nucleic Acids Res. 2015;43(1):282–94. doi: 10.1093/nar/gku1301 25505145; PubMed Central PMCID: PMC4288191.

50. Iyama T, Wilson DM 3rd., DNA repair mechanisms in dividing and non-dividing cells. DNA Repair. 2013;12(8):620–36. doi: 10.1016/j.dnarep.2013.04.015 23684800; PubMed Central PMCID: PMC3720834.

51. Scovassi AI, Prosperi E. Analysis of proliferating cell nuclear antigen (PCNA) associated with DNA. Methods in molecular biology. 2006;314:457–75. Epub 2006/05/06. doi: 10.1385/1-59259-973-7:457 16673899.

52. Balajee AS, Geard CR. Chromatin-bound PCNA complex formation triggered by DNA damage occurs independent of the ATM gene product in human cells. Nucleic Acids Res. 2001;29(6):1341–51. doi: 10.1093/nar/29.6.1341 11239001; PubMed Central PMCID: PMC29758.

53. Maugeri-Sacca M, Bartucci M, De Maria R. DNA damage repair pathways in cancer stem cells. Mol Cancer Ther. 2012;11(8):1627–36. doi: 10.1158/1535-7163.MCT-11-1040 22844074.

54. Cheung-Ong K, Giaever G, Nislow C. DNA-damaging agents in cancer chemotherapy: serendipity and chemical biology. Chem Biol. 2013;20(5):648–59. doi: 10.1016/j.chembiol.2013.04.007 23706631.

55. Pospiech H, Rytkonen AK, Syvaoja JE. The role of DNA polymerase activity in human non-homologous end joining. Nucleic Acids Res. 2001;29(15):3277–88. doi: 10.1093/nar/29.15.3277 11470886; PubMed Central PMCID: PMC55831.

56. Morgan MA, Lawrence TS. Molecular Pathways: Overcoming Radiation Resistance by Targeting DNA Damage Response Pathways. Clin Cancer Res. 2015;21(13):2898–904. Epub 2015/07/03. doi: 10.1158/1078-0432.CCR-13-3229 26133775; PubMed Central PMCID: PMC4494107.

57. Scharer OD. Nucleotide excision repair in eukaryotes. Cold Spring Harb Perspect Biol. 2013;5(10):a012609. Epub 2013/10/03. doi: 10.1101/cshperspect.a012609 24086042; PubMed Central PMCID: PMC3783044.

58. Martin LP, Hamilton TC, Schilder RJ. Platinum resistance: the role of DNA repair pathways. Clin Cancer Res. 2008;14(5):1291–5. doi: 10.1158/1078-0432.CCR-07-2238 18316546.

59. Zeman MK, Cimprich KA. Causes and consequences of replication stress. Nat Cell Biol. 2014;16(1):2–9. Epub 2013/12/25. doi: 10.1038/ncb2897 24366029; PubMed Central PMCID: PMC4354890.

60. Gary R, Ludwig DL, Cornelius HL, MacInnes MA, Park MS. The DNA repair endonuclease XPG binds to proliferating cell nuclear antigen (PCNA) and shares sequence elements with the PCNA-binding regions of FEN-1 and cyclin-dependent kinase inhibitor p21. J Biol Chem. 1997;272(39):24522–9. Epub 1997/09/26. doi: 10.1074/jbc.272.39.24522 9305916.

61. Miura M, Sasaki T. Effect of XPA gene mutations on UV-induced immunostaining of PCNA in fibroblasts from xeroderma pigmentosum group A patients. Mutat Res. 1996;364(1):51–6. Epub 1996/09/02. doi: 10.1016/0921-8777(96)00021-3 8814338.

62. Lebel M, Spillare EA, Harris CC, Leder P. The Werner syndrome gene product co-purifies with the DNA replication complex and interacts with PCNA and topoisomerase I. J Biol Chem. 1999;274(53):37795–9. Epub 1999/12/23. doi: 10.1074/jbc.274.53.37795 10608841.

63. Inoue A, Kikuchi S, Hishiki A, Shao Y, Heath R, Evison BJ, et al. A small molecule inhibitor of monoubiquitinated Proliferating Cell Nuclear Antigen (PCNA) inhibits repair of interstrand DNA cross-link, enhances DNA double strand break, and sensitizes cancer cells to cisplatin. J Biol Chem. 2014;289(10):7109–20. doi: 10.1074/jbc.M113.520429 24474685; PubMed Central PMCID: PMC3945371.


Článek vyšel v časopise

PLOS One


2019 Číslo 10

Nejčtenější v tomto čísle

Tomuto tématu se dále věnují…


Kurzy

Zvyšte si kvalifikaci online z pohodlí domova

Léčba bolesti v ordinaci praktického lékaře
nový kurz
Autoři: MUDr. PhDr. Zdeňka Nováková, Ph.D.

Revmatoidní artritida: včas a k cíli
Autoři: MUDr. Heřman Mann

Jistoty a nástrahy antikoagulační léčby aneb kardiolog - neurolog - farmakolog - nefrolog - právník diskutují
Autoři: doc. MUDr. Štěpán Havránek, Ph.D., prof. MUDr. Roman Herzig, Ph.D., doc. MUDr. Karel Urbánek, Ph.D., prim. MUDr. Jan Vachek, MUDr. et Mgr. Jolana Těšínová, Ph.D.

Léčba akutní pooperační bolesti
Autoři: doc. MUDr. Jiří Málek, CSc.

Nové antipsychotikum kariprazin v léčbě schizofrenie
Autoři: prof. MUDr. Cyril Höschl, DrSc., FRCPsych.

Všechny kurzy
Kurzy Doporučená témata Časopisy
Přihlášení
Zapomenuté heslo

Nemáte účet?  Registrujte se

Zapomenuté heslo

Zadejte e-mailovou adresu se kterou jste vytvářel(a) účet, budou Vám na ni zaslány informace k nastavení nového hesla.

Přihlášení

Nemáte účet?  Registrujte se