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Selective estrogen receptor modulators and betulinic acid act synergistically to target ERα and SP1 transcription factor dependent Pygopus expression in breast cancer
  1. Youlian R Tzenov1,
  2. Phillip Andrews1,
  3. Kim Voisey2,
  4. Luis Gai2,
  5. Beverley Carter2,
  6. Kathryn Whelan2,
  7. Catherine Popadiuk3,
  8. Kenneth R Kao1,2
  1. 1Divisions of BioMedical Science, Memorial University, St. John's, Newfoundland, Canada
  2. 2Division of Laboratory Medicine, Eastern Health, St. John's, Newfoundland, Canada
  3. 3Women's Health, Memorial University, St. John's, Newfoundland, Canada
  1. Correspondence to Dr Kenneth R Kao, Faculty of Medicine, Memorial University and Division of Anatomical Pathology, Eastern Health, 300 Prince Philip Drive, St. John's, Newfoundland, Canada A1B 3V6; kkao{at}mun.ca

Abstract

Aims 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.

  • BREAST CANCER
  • MOLECULAR BIOLOGY
  • ONCOGENES

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