title: Antisense Bcl-xl down-regulation switches the response to topoisomerase I inhibition from senescence to apoptosis in colorectal cancer cells, enhancing global cytotoxicity.
abstract: OBJECTIVE
  To identify determinants of the effect of antisense-mediated Bcl-xl down-regulation (Bcl-xl knockdown) on the response of colorectal cancer cells to SN38, the active metabolite of irinotecan, a topoisomerase I inhibitor licensed for colorectal cancer chemotherapy.
  
  
  METHODS
  Using wild-type HCT116, p53 null, Bax null, or p21/WAF1 null isogenic derivatives, we measured expression of regulators of cellular response, and associated growth arrests or apoptosis, after SN38 treatment, with or without antisense-mediated Bcl-xl knockdown.
  
  
  RESULTS
  A modified phosphorothioate antisense oligonucleotide (ISIS15999) reduced Bcl-xl protein expression by approximately 90%. SN38 induced p53, Bax, Bcl-xl, and p53-dependent p21/WAF1 protein accumulation. The Bax:Bcl-xl ratio changed little. In wild-type HCT116, but not in Bax null cells, Bcl-xl knockdown induced a shift in response from drug-induced senescence to apoptosis, and enhanced the global cytotoxicity of SN38. In p53 null or p21/WAF1 null cells marked apoptosis occurred after SN38 alone, and was additionally enhanced by Bcl-xl knockdown in p21/WAF1 null cells but not in p53 null cells.
  
  
  CONCLUSIONS
  Drug-induced senescence is associated with late relapse after therapy in transgenic models of cancer in vivo. We have shown that abolition of p21/WAF1-mediated drug-induced senescence or antisense-mediated Bcl-xl knockdown can both, independently, enhance the apoptotic response of colorectal cancer cells to SN38 in vitro. The growth arrest suppresses a p53-independent apoptotic pathway, whereas Bcl-xl induction suppresses a p53 and Bax-dependent apoptotic pathway. The combination of irinotecan and Bcl-xL antisense merits testing in models of colorectal cancer in vivo.
authors.affiliation: Cancer Research United Kingdom, Western General Hospital, Edinburgh, EH4 2XU, United Kingdom. richard.hayward@cancer.org.uk
authors.forename: Richard L
authors.initials: RL
authors.lastname: Hayward
authors.forename: Janet S
authors.initials: JS
authors.lastname: Macpherson
authors.forename: Jeff
authors.initials: J
authors.lastname: Cummings
authors.forename: Brett P
authors.initials: BP
authors.lastname: Monia
authors.forename: John F
authors.initials: JF
authors.lastname: Smyth
authors.forename: Duncan I
authors.initials: DI
authors.lastname: Jodrell
mesh_terms: D017304:Annexin A5 / Q000494:pharmacology; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D017209:Apoptosis; D002166:Camptothecin / Q000031:analogs & derivatives* / Q000494:pharmacology; D002453:Cell Cycle; D002455:Cell Division; D045744:Cell Line, Tumor; D016922:Cellular Senescence; D015179:Colorectal Neoplasms / Q000473:pathology; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005456:Fluorescent Dyes / Q000494:pharmacology; D016158:Genes, p53 / Q000235:genetics; D006801:Humans; D015151:Immunoblotting; D000077146:Irinotecan; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D011518:Proto-Oncogene Proteins / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics* / Q000378:metabolism; D013997:Time Factors; D059004:Topoisomerase I Inhibitors; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: D017304:Annexin A5; D000972:Antineoplastic Agents, Phytogenic; C494826:BAX protein, human; C494811:BCL2L1 protein, human; C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D016376:Oligonucleotides, Antisense; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D059004:Topoisomerase I Inhibitors; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; D000077146:Irinotecan; D001616:beta-Galactosidase; D002166:Camptothecin