title: Selective estrogen receptor modulators and betulinic acid act synergistically to target ERα and SP1 transcription factor dependent Pygopus expression in breast cancer.
abstract: OBJECTIVE
  Estrogen and progesterone hormone receptor (ER and PR) expression in invasive breast cancer predicts response to hormone disruptive therapy. Pygopus2 (hPYGO2) encodes a chromatin remodelling protein important for breast cancer growth and cell cycle progression. The aims of this study were to determine the mechanism of expression of hPYGO2 in breast cancer and to examine how this expression is affected therapeutically.
  
  
  METHODS
  hPYGO2 and ER protein expression was examined in a breast tumour microarray by immunohistochemistry. hPYGO2 RNA and protein expression was examined in ER+ and ER- breast cancer cell lines in the presence of selective estrogen hormone receptor modulator drugs and the specificity protein-1 (SP1) inhibitor, betulinic acid (BA). The effects of these drugs on the ability for ER and SP1 to bind the hPYGO2 promoter and affect cell cycle progression were studied using chromatin immunoprecipitation assays.
  
  
  RESULTS
  hPYGO2 was expressed in seven of eight lines and in nuclei of 98% of 65 breast tumours, including 3 Ductal carcinoma in situ and 62 invasive specimens representing ER-negative (22%) and ER-positive (78%) cases. Treatment with either 4-Hydroxytamoxifen (OHT) or fulvestrant reduced hPYGO2 mRNA 10-fold and protein 5-10-fold within 4 h. Promoter analysis indicated an ER/SP1 binding site at nt -225 to -531 of hPYGO2. SP1 RNA interference and BA reduced hPYGO2 protein and RNA expression by fivefold in both ER- and ER+ cells. Further attenuation was achieved by combining BA and 4-OHT resulting in eightfold reduction in cell growth.
  
  
  CONCLUSIONS
  Our findings reveal a mechanistic link between hormone signalling and the growth transcriptional programme. The activation of its expression by ERα and/or SP1 suggests hPYGO2 as a theranostic target for hormone therapy responsive and refractory breast cancer.
authors.affiliation: Divisions of BioMedical Science, Memorial University, St. John's, Newfoundland, Canada.
authors.forename: Youlian R
authors.initials: YR
authors.lastname: Tzenov
authors.affiliation: Divisions of BioMedical Science, Memorial University, St. John's, Newfoundland, Canada.
authors.forename: Phillip
authors.initials: P
authors.lastname: Andrews
authors.affiliation: Division of Laboratory Medicine, Eastern Health, St. John's, Newfoundland, Canada.
authors.forename: Kim
authors.initials: K
authors.lastname: Voisey
authors.affiliation: Division of Laboratory Medicine, Eastern Health, St. John's, Newfoundland, Canada.
authors.forename: Luis
authors.initials: L
authors.lastname: Gai
authors.affiliation: Division of Laboratory Medicine, Eastern Health, St. John's, Newfoundland, Canada.
authors.forename: Beverley
authors.initials: B
authors.lastname: Carter
authors.affiliation: Division of Laboratory Medicine, Eastern Health, St. John's, Newfoundland, Canada.
authors.forename: Kathryn
authors.initials: K
authors.lastname: Whelan
authors.affiliation: Women's Health, Memorial University, St. John's, Newfoundland, Canada.
authors.forename: Catherine
authors.initials: C
authors.lastname: Popadiuk
authors.affiliation: Divisions of BioMedical Science, Memorial University, St. John's, Newfoundland, Canada Division of Laboratory Medicine, Eastern Health, St. John's, Newfoundland, Canada.
authors.forename: Kenneth R
authors.initials: KR
authors.lastname: Kao
mesh_terms: D000970:Antineoplastic Agents / Q000627:therapeutic use; D001943:Breast Neoplasms / Q000378:metabolism*; D002285:Carcinoma, Intraductal, Noninfiltrating / Q000378:metabolism*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004958:Estradiol / Q000031:analogs & derivatives / Q000627:therapeutic use; D047628:Estrogen Receptor alpha / Q000235:genetics / Q000378:metabolism*; D005260:Female; D000077267:Fulvestrant; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D006915:Hydroxytestosterones / Q000627:therapeutic use; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D046228:Microarray Analysis; D053978:Pentacyclic Triterpenes; D011401:Promoter Regions, Genetic / Q000235:genetics; D020845:Selective Estrogen Receptor Modulators / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D016329:Sp1 Transcription Factor / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D013629:Tamoxifen / Q000031:analogs & derivatives / Q000627:therapeutic use; D014315:Triterpenes / Q000627:therapeutic use; D000094062:Betulinic Acid
chemical_list: D000970:Antineoplastic Agents; C506487:ESR1 protein, human; D047628:Estrogen Receptor alpha; D006915:Hydroxytestosterones; D047908:Intracellular Signaling Peptides and Proteins; C453401:PYGO2 protein, human; D053978:Pentacyclic Triterpenes; D020845:Selective Estrogen Receptor Modulators; D016329:Sp1 Transcription Factor; C108209:SP1 protein, human; D014315:Triterpenes; D013629:Tamoxifen; C016601:afimoxifene; C032278:4,17 beta-dihydroxy-4-androstene-3-one; D000077267:Fulvestrant; D004958:Estradiol; D000094062:Betulinic Acid
keywords: BREAST CANCER; MOLECULAR BIOLOGY; ONCOGENES