title: Rad51 and BRCA2--New molecular targets for sensitizing glioma cells to alkylating anticancer drugs.
abstract: First line chemotherapeutics for brain tumors (malignant gliomas) are alkylating agents such as temozolomide and nimustine. Despite growing knowledge of how these agents work, patients suffering from this malignancy still face a dismal prognosis. Alkylating agents target DNA, forming the killing lesion O(6)-alkylguanine, which is converted into DNA double-strand breaks (DSBs) that trigger apoptosis. Here we assessed whether inhibiting repair of DSBs by homologous recombination (HR) or non-homologous end joining (NHEJ) is a reasonable strategy for sensitizing glioma cells to alkylating agents. For down-regulation of HR in glioma cells, we used an interference RNA (iRNA) approach targeting Rad51 and BRCA2, and for NHEJ we employed the DNA-PK inhibitor NU7026. We also assessed whether inhibition of poly(ADP)ribosyltransferase (PARP) by olaparib would enhance the killing effect. The data show that knockdown of Rad51 or BRCA2 greatly sensitizes cells to DSBs and the induction of cell death following temozolomide and nimustine (ACNU). It did not sensitize to ionizing radiation (IR). The expression of O(6)-methylguanine-DNA methyltransferase (MGMT) abolished all these effects, indicating that O(6)-alkylguanine induced by these drugs is the primary lesion responsible for the formation of DSBs and increased sensitivity of glioma cells following knockdown of Rad51 and BRCA2. Inhibition of DNA-PK only slightly sensitized to temozolomide whereas a significant effect was observed with IR. A triple strategy including siRNA and the PARP inhibitor olaparib further improved the killing effect of temozolomide. The data provides evidence that down-regulation of Rad51 or BRCA2 is a reasonable strategy for sensitizing glioma cells to killing by O(6)-alkylating anti-cancer drugs. The data also provide proof of principle that a triple strategy involving down-regulation of HR, PARP inhibition and MGMT depletion may greatly enhance the therapeutic effect of temozolomide.
authors.affiliation: Institute for Toxicology, Medical Center of the Johannes Gutenberg University Mainz, Mainz, Germany.
authors.forename: Steve
authors.initials: S
authors.lastname: Quiros
authors.forename: Wynand Paul
authors.initials: WP
authors.lastname: Roos
authors.forename: Bernd
authors.initials: B
authors.lastname: Kaina
mesh_terms: D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology / Q000627:therapeutic use*; D017209:Apoptosis / Q000187:drug effects; D024682:BRCA2 Protein / Q000187:drug effects*; D001483:Base Sequence; D001932:Brain Neoplasms / Q000188:drug therapy* / Q000473:pathology; D045744:Cell Line, Tumor; D002867:Chromones / Q000494:pharmacology; D004249:DNA Damage; D005434:Flow Cytometry; D005910:Glioma / Q000188:drug therapy* / Q000473:pathology; D059765:Homologous Recombination; D006801:Humans; D008856:Microscopy, Fluorescence; D009025:Morpholines / Q000494:pharmacology; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D034741:RNA, Small Interfering; D051135:Rad51 Recombinase / Q000187:drug effects*
chemical_list: C479235:2-(morpholin-4-yl)benzo(h)chromen-4-one; D018906:Antineoplastic Agents, Alkylating; D024682:BRCA2 Protein; D002867:Chromones; D009025:Morpholines; D047428:Protein Kinase Inhibitors; D034741:RNA, Small Interfering; C495153:RAD51 protein, human; D051135:Rad51 Recombinase
references.citation: Cell Res. 2008 Jan;18(1):114-24
references.pmid: 18166980
references.citation: Oncogene. 2007 Jan 11;26(2):186-97
references.pmid: 16819506
references.citation: Genes Dev. 2007 Dec 15;21(24):3342-55
references.pmid: 18079180
references.citation: DNA Repair (Amst). 2010 Oct 5;9(10):1050-63
references.pmid: 20708982
references.citation: Mutat Res. 1990 Jul;231(1):11-30
references.pmid: 2195323
references.citation: Oncol Res. 1995;7(1):49-55
references.pmid: 7549044
references.citation: Nucleic Acids Res. 2007;35(11):3535-50
references.pmid: 17483520
references.citation: Cancer Res. 2007 Dec 15;67(24):11886-95
references.pmid: 18089819
references.citation: J Neurochem. 2006 Feb;96(3):766-76
references.pmid: 16405512
references.citation: Nature. 2010 Oct 7;467(7316):678-83
references.pmid: 20729832
references.citation: Int J Cancer. 2010 Nov 1;127(9):2106-18
references.pmid: 20131314
references.citation: N Engl J Med. 2005 Mar 10;352(10):987-96
references.pmid: 15758009
references.citation: EMBO J. 2006 Jan 11;25(1):222-31
references.pmid: 16395335
references.citation: Annu Rev Biochem. 2010;79:181-211
references.pmid: 20192759
references.citation: Cell Cycle. 2010 Jan 1;9(1):168-78
references.pmid: 20016283
references.citation: Annu Rev Biochem. 2008;77:229-57
references.pmid: 18275380
references.citation: Carcinogenesis. 1991 Oct;12(10):1857-67
references.pmid: 1657427
references.citation: Brain Res. 2010 Jun 18;1338:112-21
references.pmid: 20307511
references.citation: Bioessays. 1994 Nov;16(11):833-9
references.pmid: 7840761
references.citation: DNA Repair (Amst). 2009 Jan 1;8(1):72-86
references.pmid: 18840549
references.citation: Hum Mol Genet. 2011 Apr 15;20(R1):R28-41
references.pmid: 21429918
references.citation: Int J Cancer. 1998 Dec 18;79(6):640-4
references.pmid: 9842975
references.citation: N Engl J Med. 2005 Mar 10;352(10):997-1003
references.pmid: 15758010
references.citation: Nat Struct Mol Biol. 2010 Oct;17(10):1260-2
references.pmid: 20729859
references.citation: Nature. 2005 Apr 14;434(7035):913-7
references.pmid: 15829966
references.citation: Br J Cancer. 2006 Feb 13;94(3):407-26
references.pmid: 16434996
references.citation: J Neurooncol. 2007 Mar;82(1):85-9
references.pmid: 17031555
references.citation: Blood. 2004 Jun 15;103(12):4659-65
references.pmid: 15010369
references.citation: J Neurooncol. 2008 Aug;89(1):27-35
references.pmid: 18415044
references.citation: DNA Repair (Amst). 2007 Aug 1;6(8):1079-99
references.pmid: 17485253
references.citation: Cell Mol Life Sci. 2010 Nov;67(21):3663-81
references.pmid: 20717836
references.citation: Cancer Res. 2000 Oct 15;60(20):5815-24
references.pmid: 11059778
references.citation: J Biol Chem. 1998 Mar 6;273(10):5858-68
references.pmid: 9488723
references.citation: Cell Res. 2008 Jan;18(1):99-113
references.pmid: 18166982