title: Inhibition of Sp1 activity by a decoy PNA-DNA chimera prevents urokinase receptor expression and migration of breast cancer cells.
abstract: Sp1 regulates the activation of many genes involved in tumor growth, apoptosis, and angiogenesis. We have previously shown the involvement of Sp1 in the up-regulation of urokinase receptor (uPAR) expression, a key molecule in tumor invasion and metastasis. Here, we investigated whether a marked down-regulation of Sp1 activity may inhibit uPAR expression and migration ability of MDA-MB-231 breast cancer cells. To this end, we tested the decoy ability of a novel peptide nucleic acid (PNA)-DNA chimera which carries a central DNA strand, containing Sp1-binding sequence, covalently linked to two PNA fragments at both ends (PNA-DNA-PNA, PDP). The chimera was synthesized, annealed with complementary DNA (PDP-DNA), and then tested for its ability to bind Sp1 both in vitro and in living MDA-MB-231 breast cancer cells in the presence of urokinase (uPA). This PDP-DNA decoy molecule efficiently competes for the binding to endogenous Sp1 in nuclear extracts, and upon transfection with liposomal vectors, causes a marked decrease of available Sp1 in both untreated and uPA-treated MDA-MB-231 cells. Accordingly, both uPA-dependent enhancement of uPAR expression and cell migration were strongly reduced in transfected cells. Interestingly, a detectable inhibitory effect is also observed in breast cancer cells exposed to PDP-DNA in the absence of transfection reagents. Finally, the inhibitory effect of PDP-DNA appeared to be stronger than that observed with oligonucleotides carrying Sp1 consensus sequence. Our findings show that this novel PNA-DNA chimera, containing Sp1 consensus sequence, effectively inhibits Sp1 activity, uPAR expression, and motility of breast cancer cells indicating its potential therapeutic use to prevent tumor dissemination.
authors.affiliation: Istituto di Biostrutture e Bioimmagini, Consiglio Nazionale delle Ricerche, Via S. Pansini 5, 80131 Naples, Italy.
authors.forename: Antonella
authors.initials: A
authors.lastname: Zannetti
authors.forename: Silvana
authors.initials: S
authors.lastname: Del Vecchio
authors.forename: Alessandra
authors.initials: A
authors.lastname: Romanelli
authors.forename: Stefania
authors.initials: S
authors.lastname: Scala
authors.forename: Michele
authors.initials: M
authors.lastname: Saviano
authors.forename: Gaetano
authors.initials: G
authors.lastname: Cali'
authors.forename: M Patrizia
authors.initials: MP
authors.lastname: Stoppelli
authors.forename: Carlo
authors.initials: C
authors.lastname: Pedone
authors.forename: Marco
authors.initials: M
authors.lastname: Salvatore
mesh_terms: D001943:Breast Neoplasms / Q000737:chemistry / Q000188:drug therapy* / Q000473:pathology; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D004247:DNA / Q000494:pharmacology*; D005260:Female; D006801:Humans; D020135:Peptide Nucleic Acids / Q000494:pharmacology*; D011956:Receptors, Cell Surface / Q000032:analysis / Q000037:antagonists & inhibitors*; D055293:Receptors, Urokinase Plasminogen Activator; D016329:Sp1 Transcription Factor / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: C524898:PLAUR protein, human; D020135:Peptide Nucleic Acids; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D016329:Sp1 Transcription Factor; D004247:DNA