title: Estrogen Receptor β Agonists Differentially Affect the Growth of Human Melanoma Cell Lines.
abstract: BACKGROUND
  Cutaneous melanoma is an aggressive malignancy; its incidence is increasing worldwide and its prognosis remains poor. Clinical observations indicate that estrogen receptor β (ERβ) is expressed in melanoma tissues and its expression decreases with tumor progression, suggesting its tumor suppressive function. These experiments were performed to investigate the effects of ERβ activation on melanoma cell growth.
  
  
  RESULTS
  Protein expression was analyzed by Western blot and immunofluorescence assays. Cell proliferation was assessed by counting the cells by hemocytometer. ERβ transcriptional activity was evaluated by gene reporter assay. Global DNA methylation was analyzed by restriction enzyme assay and ERβ isoforms were identified by qRT-PCR. We demonstrated that ERβ is expressed in a panel of human melanoma cell lines (BLM, WM115, A375, WM1552). In BLM (NRAS-mutant) cells, ERβ agonists significantly and specifically inhibited cell proliferation. ERβ activation triggered its cytoplasmic-to-nuclear translocation and transcriptional activity. Moreover, the antiproliferative activity of ERβ agonists was associated with an altered expression of G1-S transition-related proteins. In these cells, global DNA was found to be hypomethylated when compared to normal melanocytes; this DNA hypomethylation status was reverted by ERβ activation. ERβ agonists also decreased the proliferation of WM115 (BRAF V600D-mutant) cells, while they failed to reduce the growth of A375 and WM1552 (BRAF V600E-mutant) cells. Finally, we could observe that ERβ isoforms are expressed at different levels in the various cell lines. Specific oncogenic mutations or differential expression of receptor isoforms might be responsible for the different responses of cell lines to ERβ agonists.
  
  
  CONCLUSIONS
  Our results demonstrate that ERβ is expressed in melanoma cell lines and that ERβ agonists differentially regulate the proliferation of these cells. These data confirm the notion that melanoma is a heterogeneous tumor and that genetic profiling is mandatory for the development of effective personalized therapeutic approaches for melanoma patients.
authors.affiliation: Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, Milano, Italy.
authors.forename: Monica
authors.initials: M
authors.lastname: Marzagalli
authors.affiliation: Department of Medical Biotechnologies and Translational Medicine, Università degli Studi di Milano, Milano, Italy.
authors.forename: Lavinia
authors.initials: L
authors.lastname: Casati
authors.affiliation: Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, Milano, Italy.
authors.forename: Roberta M
authors.initials: RM
authors.lastname: Moretti
authors.affiliation: Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, Milano, Italy.
authors.forename: Marina
authors.initials: M
authors.lastname: Montagnani Marelli
authors.affiliation: Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, Milano, Italy.
authors.forename: Patrizia
authors.initials: P
authors.lastname: Limonta
mesh_terms: D018797:Cell Cycle Proteins / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects*; D045744:Cell Line, Tumor; D019175:DNA Methylation; D047629:Estrogen Receptor beta / Q000819:agonists*; D004967:Estrogens / Q000494:pharmacology*; D006801:Humans; D008545:Melanoma / Q000378:metabolism / Q000473:pathology*
chemical_list: D018797:Cell Cycle Proteins; D047629:Estrogen Receptor beta; D004967:Estrogens