Imbalancing DNA BER to enhance Ovarian Tumor Sensitivity

Summary

Principal Investigator: Mark Kelley
Abstract: The overall significance of this project relates to the ability to imbalance the DNA base excision repair (BER) pathway in ovarian tumor ceils, increasing their sensitivity to chemotherapeutic and ionizing radiation (IR) agents. We will attempt to accomplish this goal using mutants of the human apurinic/apyrimidinic endonuclease (APE1) enzyme, overexpression of N-methylpurine DNA glycosylase (MPG), both targeted to the nucleus and mitochondria, as well as small (short) interfering RNA (siRNA) for APE1. We will also utilize folic acid-derivatized liposomes and adenoviral targeting along with tumor specific promoter expression using the human telomerase reverste transcriptase (hTERT) promoter in both cell lines and an NOD/SCID animal model to develop the usefulness of this approach. Hypothesis: Overexpression of MPG in the nucleus and/or mitochondrial compartments, altered human APE1 proteins (dominant-negative), or siRNA for APE1 either independently, or in various combinations will enhance ovarian cancer cells to standard or decreased levels of commonly used chemotherapeutic agents (e.g. alkylators) and/or IR. The Specific Aims are: Specific Aim 1: This first aim includes determining the effectiveness of overexpressing MPG or dominant-negative APE1 in multiple ovarian cancer lines and evaluating tumor cell response to chemotherapeutic and IR treatment. This includes both nuclear and mitochondrial targeting of the MPG enzyme and overexpression and the knockdown of APE1 with siRNA. Specific Aim 2: Determine the effects of co-overexpression of nucMPG, mitoMPG and nuclMPG+mitoMPG, nucMPG+APE1 mutant, mitoMPG+APE1 mutant and nucMPG or mitoMPG and APEI-siRNA. We will monitor whether combined expression enhances the tumor cell killing effect of chemotherapeutic agents or IR. Specific Aim 3: Constructs using the hTERT promoter will be used in ovarian cancer cell lines in both plasmid (folic acid-derivatized liposome) and adenoviral based delivery systems for tumor specific expression studies using best candidate APE1 mutants, nuc- or mitoMPG, or siRNA as determined by the results in Aims 1-2. Specific Aim 4: Determine in vivo chemo- and radiosensitivity due to the expression of the various constructs of APE1 mutants, or nuc-/mitoMPG as well as APEI-siRNA in NOD/SCID mice. Adenoviral constructs with the hTERT promoter as well as folic acid-derivatized liposomes containing selected genes from the first three aims will be used with xenograft NOD/SCID mice. If successful, we feel these studies will create very effective reagents in a therapeutic gene transfer/therapy setting in the clinic, as well as shed light on the role of both nuclear and mitochondrial BER in cancer cells.
Funding Period: 2004-05-01 - 2010-04-30
more information: NIH RePORT

Top Publications

  1. ncbi The DNA base excision repair protein Ape1/Ref-1 as a therapeutic and chemopreventive target
    Melissa L Fishel
    Department of Pediatrics Section of Hematology Oncology, Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN 46202, United States
    Mol Aspects Med 28:375-95. 2007
  2. pmc Semi-automated high-throughput fluorescent intercalator displacement-based discovery of cytotoxic DNA binding agents from a large compound library
    LaTeca S Glass
    Department of Chemistry and Chemical Biology, Purdue School of Science, Indiana University Purdue University Indianapolis, Indianapolis, IN 46202, USA
    Bioorg Med Chem Lett 20:1685-8. 2010
  3. pmc Redox regulation of DNA repair: implications for human health and cancer therapeutic development
    Meihua Luo
    Department of Pediatrics, Herman B Wells Center for Pediatric Research, Indiana University, Indiana, USA
    Antioxid Redox Signal 12:1247-69. 2010
  4. pmc Inhibition of the redox function of APE1/Ref-1 in myeloid leukemia cell lines results in a hypersensitive response to retinoic acid-induced differentiation and apoptosis
    Melissa L Fishel
    Department of Pediatrics Section of Hematology Oncology, Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, 980 W Walnut, Indianapolis, IN 46202, USA
    Exp Hematol 38:1178-88. 2010
  5. pmc Novel small-molecule inhibitor of apurinic/apyrimidinic endonuclease 1 blocks proliferation and reduces viability of glioblastoma cells
    Aditi Bapat
    Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis, Indiana, USA
    J Pharmacol Exp Ther 334:988-98. 2010
  6. pmc Design and synthesis of novel quinone inhibitors targeted to the redox function of apurinic/apyrimidinic endonuclease 1/redox enhancing factor-1 (Ape1/ref-1)
    Rodney L Nyland
    Department of Medicinal Chemistry and Molecular Pharmacology, Cancer Center, Purdue University, West Lafayette, Indiana 47907, USA
    J Med Chem 53:1200-10. 2010
  7. pmc Reduced expression of DNA repair and redox signaling protein APE1/Ref-1 impairs human pancreatic cancer cell survival, proliferation, and cell cycle progression
    Yanlin Jiang
    Department of Pediatrics, Herman B Wells Center for Pediatric Research, Walnut, Indianapolis 46202, USA
    Cancer Invest 28:885-95. 2010
  8. pmc Knock-in reconstitution studies reveal an unexpected role of Cys-65 in regulating APE1/Ref-1 subcellular trafficking and function
    Carlo Vascotto
    Department of Medical and Biological Sciences, University of Udine, 33100 Udine, Italy
    Mol Biol Cell 22:3887-901. 2011
  9. pmc Base excision repair apurinic/apyrimidinic endonucleases in apicomplexan parasite Toxoplasma gondii
    David O Onyango
    Department of Pharmacology and Toxicology, Indiana University School of Medicine, Indianapolis, 46202, United States
    DNA Repair (Amst) 10:466-75. 2011
  10. pmc Mad2 haploinsufficiency protects hematopoietic progenitor cells subjected to cell-cycle stress in vivo and to inhibition of redox function of Ape1/Ref-1 in vitro
    Sara L Rohrabaugh
    Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, IN 46202, USA
    Exp Hematol 39:415-23. 2011

Detail Information

Publications29

  1. ncbi The DNA base excision repair protein Ape1/Ref-1 as a therapeutic and chemopreventive target
    Melissa L Fishel
    Department of Pediatrics Section of Hematology Oncology, Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN 46202, United States
    Mol Aspects Med 28:375-95. 2007
    ..In this review, we will provide an overview of Ape1/Ref-1's activities and explore the potential of this protein as a target in cancer treatment as well as its role in chemoprevention...
  2. pmc Semi-automated high-throughput fluorescent intercalator displacement-based discovery of cytotoxic DNA binding agents from a large compound library
    LaTeca S Glass
    Department of Chemistry and Chemical Biology, Purdue School of Science, Indiana University Purdue University Indianapolis, Indianapolis, IN 46202, USA
    Bioorg Med Chem Lett 20:1685-8. 2010
    ..In addition, the general screening strategy described may find broader impact toward the rapid discovery of DNA targeted agents with biological activity...
  3. pmc Redox regulation of DNA repair: implications for human health and cancer therapeutic development
    Meihua Luo
    Department of Pediatrics, Herman B Wells Center for Pediatric Research, Indiana University, Indiana, USA
    Antioxid Redox Signal 12:1247-69. 2010
    ..Finally, we consider the potential for chemotherapeutic development through the modulation of APE1's redox activity and its impact on DNA repair...
  4. pmc Inhibition of the redox function of APE1/Ref-1 in myeloid leukemia cell lines results in a hypersensitive response to retinoic acid-induced differentiation and apoptosis
    Melissa L Fishel
    Department of Pediatrics Section of Hematology Oncology, Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, 980 W Walnut, Indianapolis, IN 46202, USA
    Exp Hematol 38:1178-88. 2010
    ....
  5. pmc Novel small-molecule inhibitor of apurinic/apyrimidinic endonuclease 1 blocks proliferation and reduces viability of glioblastoma cells
    Aditi Bapat
    Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis, Indiana, USA
    J Pharmacol Exp Ther 334:988-98. 2010
    ..AR03 is a novel small-molecule inhibitor of Ape1, which may have potential as an oncotherapeutic drug for treating glioblastoma and other cancers...
  6. pmc Design and synthesis of novel quinone inhibitors targeted to the redox function of apurinic/apyrimidinic endonuclease 1/redox enhancing factor-1 (Ape1/ref-1)
    Rodney L Nyland
    Department of Medicinal Chemistry and Molecular Pharmacology, Cancer Center, Purdue University, West Lafayette, Indiana 47907, USA
    J Med Chem 53:1200-10. 2010
    ..Most of the naphthoquinones were low micromolar inhibitors of Ape1 redox activity, and the most potent analogues inhibited tumor cell growth with IC(50) values in the 10-20 microM range...
  7. pmc Reduced expression of DNA repair and redox signaling protein APE1/Ref-1 impairs human pancreatic cancer cell survival, proliferation, and cell cycle progression
    Yanlin Jiang
    Department of Pediatrics, Herman B Wells Center for Pediatric Research, Walnut, Indianapolis 46202, USA
    Cancer Invest 28:885-95. 2010
    ..Endogenous cell cycle inhibitors increase when APE1/ Ref-1 is reduced, demonstrating its importance to proliferation and growth of pancreatic cancer...
  8. pmc Knock-in reconstitution studies reveal an unexpected role of Cys-65 in regulating APE1/Ref-1 subcellular trafficking and function
    Carlo Vascotto
    Department of Medical and Biological Sciences, University of Udine, 33100 Udine, Italy
    Mol Biol Cell 22:3887-901. 2011
    ....
  9. pmc Base excision repair apurinic/apyrimidinic endonucleases in apicomplexan parasite Toxoplasma gondii
    David O Onyango
    Department of Pharmacology and Toxicology, Indiana University School of Medicine, Indianapolis, 46202, United States
    DNA Repair (Amst) 10:466-75. 2011
    ..The importance of TgAPN and the fact that humans lack any observable APN family activity highlights TgAPN as a promising candidate for drug development to treat toxoplasmosis...
  10. pmc Mad2 haploinsufficiency protects hematopoietic progenitor cells subjected to cell-cycle stress in vivo and to inhibition of redox function of Ape1/Ref-1 in vitro
    Sara L Rohrabaugh
    Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, IN 46202, USA
    Exp Hematol 39:415-23. 2011
    ..We evaluated effects of Mad2 haploinsufficiency on hematopoietic stem (HSC) and progenitor (HPC) function in response to stress...
  11. pmc Apurinic/Apyrimidinic endonuclease 1 regulates inflammatory response in macrophages
    Andrej Jedinak
    Cancer Research Laboratory, Methodist Research Institute, Indiana University Health, 1800 N Capitol Ave, E504, Indianapolis, IN 46202, USA
    Anticancer Res 31:379-85. 2011
    ..In conclusion, pharmacological inhibition of APE1 by E3330 suppresses inflammatory response in activated macrophages and can be considered as a novel therapeutic strategy for the inhibition of tumor-associated macrophages...
  12. pmc Functional analysis of novel analogues of E3330 that block the redox signaling activity of the multifunctional AP endonuclease/redox signaling enzyme APE1/Ref-1
    Mark R Kelley
    Department of Pediatrics Section of Hematology Oncology, Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, 980 West Walnut Street, Indianapolis, IN 46202, USA
    Antioxid Redox Signal 14:1387-401. 2011
    ..This progress in synthesizing and isolating biologically active novel E3330 analogues that effectively inhibit the APE1 redox function validates the utility of further translational anticancer therapeutic development...
  13. pmc Inhibition of APE1/Ref-1 redox activity with APX3330 blocks retinal angiogenesis in vitro and in vivo
    Aihua Jiang
    Cardiovascular Institute, Beth Israel Deaconess Medical Center, 3 Blackfan Circle, Boston, MA 02215, United States
    Vision Res 51:93-100. 2011
    ..Thus, APE1/Ref-1 may have potential as a therapeutic target for treating neovascular age-related macular degeneration and other neovascular diseases...
  14. pmc Role of APE1 in differentiated neuroblastoma SH-SY5Y cells in response to oxidative stress: use of APE1 small molecule inhibitors to delineate APE1 functions
    Yanlin Jiang
    Department of Pediatrics Section of Hematology Oncology, Herman B Wells Center for Pediatric Research, United States
    DNA Repair (Amst) 8:1273-82. 2009
    ..Our results demonstrate that the DNA repair function of APE1 contributes to the survival of nondividing post-mitotic cells following oxidative DNA damage...
  15. pmc Role of Ape1 and base excision repair in the radioresponse and heat-radiosensitization of HeLa Cells
    Christopher N Batuello
    Indiana University, Department of Radiation Oncology, Indianapolis, Indiana 46202, USA
    Anticancer Res 29:1319-25. 2009
    ..We determined whether base excision repair (BER) is involved in heat-radiosensitization and report novel findings that provide insight regarding the role of BER in the radiation response of HeLa cells...
  16. pmc Going ape as an approach to cancer therapeutics
    Aditi Bapat
    Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis, Indiana, USA
    Antioxid Redox Signal 11:651-68. 2009
    ..Therefore, selective inhibition of Ape1's DNA repair activity is a promising avenue to develop novel cancer therapeutics...
  17. ncbi Manipulation of base excision repair to sensitize ovarian cancer cells to alkylating agent temozolomide
    Melissa L Fishel
    Department of Pediatrics Section of Hematology Oncology, Herman B Wells Center for Pediatric Research, Indianapolis, Indiana 46202, USA
    Clin Cancer Res 13:260-7. 2007
    ....
  18. ncbi DNA repair in neurons: so if they don't divide what's to repair?
    Melissa L Fishel
    Department of Pediatrics, Section of Hematology Oncology, Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, 1044 W Walnut, Room 302C, Indianapolis, IN 46202, USA
    Mutat Res 614:24-36. 2007
    ..Studies of DNA repair may help our understanding of how those cells that are not dividing could succumb to neurotoxicity with the clinical manifestations discussed in the following article...
  19. ncbi Repression of cyclin D1 as a target for germ cell tumors
    Sarah J Freemantle
    Department of Pharmacology and Toxicology, Dartmouth Medical School, HB 7650, Hanover, NH 03755, USA
    Int J Oncol 30:333-40. 2007
    ..This inhibited proliferation in RA and cisplatin sensitive and resistant EC cells. Taken together, these findings implicate cyclin D1 targeting agents for the treatment of GCTs...
  20. ncbi Ape1 regulates hematopoietic differentiation of embryonic stem cells through its redox functional domain
    Gang Ming Zou
    Department of Pediatrics Section of Hematology Oncology, Indiana University School of Medicine, Indianapolis 46202, USA
    Blood 109:1917-22. 2007
    ..In summary, these data indicate Ape1 is required in normal embryonic hematopoiesis and that the redox function, but not the repair endonuclease activity, of Ape1 is critical in normal embryonic hematopoietic development...
  21. pmc Enhancement of cisplatin [cis-diammine dichloroplatinum (II)] cytotoxicity by O6-benzylguanine involves endoplasmic reticulum stress
    Cara A Rabik
    Department of Molecular Genetics and Cell Biology, University of Chicago, Chicago, Illinois 60637, USA
    J Pharmacol Exp Ther 327:442-52. 2008
    ..These data indicate GADD153 up-regulation plays an important role in BG-enhanced cisplatin cytotoxicity and apoptosis...
  22. pmc Knockdown of the DNA repair and redox signaling protein Ape1/Ref-1 blocks ovarian cancer cell and tumor growth
    Melissa L Fishel
    Department of Pediatrics Section of Hematology Oncology, Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN 46202, USA
    DNA Repair (Amst) 7:177-86. 2008
    ..Ape1's role in DNA repair and redox signaling is important to our basic understanding of ovarian cancer cell growth and these findings strongly support Ape1 as a therapeutic target...
  23. pmc Role of the multifunctional DNA repair and redox signaling protein Ape1/Ref-1 in cancer and endothelial cells: small-molecule inhibition of the redox function of Ape1
    Meihua Luo
    Department of Pediatrics Section of Hematology Oncology, Herman B Wells Center for Pediatric Research, Indianapolis, Indiana 46202, USA
    Antioxid Redox Signal 10:1853-67. 2008
    ..Here we discuss small-molecule inhibition of Ape1 redox and its effect on both cancer and endothelial cells...
  24. pmc Implications of apurinic/apyrimidinic endonuclease in reactive oxygen signaling response after cisplatin treatment of dorsal root ganglion neurons
    Yanlin Jiang
    Department of Pediatrics, Section of Hematology Oncology, Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA
    Cancer Res 68:6425-34. 2008
    ....
  25. pmc DNA repair proteins as molecular targets for cancer therapeutics
    Mark R Kelley
    Department of Pediatrics, Section of Hematology Oncology, Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, 1044 W Walnut St R4 W302C, Indianapolis, IN 46202, USA
    Anticancer Agents Med Chem 8:417-25. 2008
    ....
  26. pmc Evolution of the redox function in mammalian apurinic/apyrimidinic endonuclease
    M M Georgiadis
    Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, 635 Barnhill Drive, Indianapolis, IN 46202 5122, United States
    Mutat Res 643:54-63. 2008
    ..9 and 2.3A, respectively. Our results provide new insights on the redox function and highlight a dramatic gain-of-function activity for Ape1 in mammals not found in non-mammalian vertebrates or lower organisms...
  27. pmc Small-molecule inhibitors of proteins involved in base excision repair potentiate the anti-tumorigenic effect of existing chemotherapeutics and irradiation
    April M Reed
    Department of Pediatrics, Section of Hematology Oncology, Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN 46202, USA
    Future Oncol 5:713-26. 2009
    ....
  28. pmc The many functions of APE1/Ref-1: not only a DNA repair enzyme
    Gianluca Tell
    Department of Biomedical Sciences and Technologies, University of Udine, Udine, Italy
    Antioxid Redox Signal 11:601-20. 2009
    ....
  29. pmc APE1/Ref-1 role in redox signaling: translational applications of targeting the redox function of the DNA repair/redox protein APE1/Ref-1
    Mark R Kelley
    Department of Pediatrics Section of Hematology Oncology, Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN 46202, USA
    Curr Mol Pharmacol 5:36-53. 2012
    ..It also discusses APE1's altered expression in many cancers and the therapeutic potential of selective inhibition of redox regulation, which is the subject of intense preclinical studies...