Epz005687

The full length fragments of FOXF1 coding sequences and PVT1 gene sequences were amplified by PCR and constructed into pcDNA3.1 vector (Invitrogen) to generate pcDNA-FOXF1 and pcDNA-PVT1 (PVT1) plasmid. Small interference RNAs (siRNAs) of PVT1 (si-PVT1#1, si-PVT1#2, si-PVT1#3), FOXF1 (si-FOXF1) and EZH2 (si-EZH2) along with the scramble control (si-con) were synthesized by GenePharma Co., Ltd (Shanghai, China). All these siRNAs or plasmids were transfected into breast cancer cells using lipofectamine 2000 reagent (Invitrogen) following the manufacturer's protocol. EZH2 inhibitor EPZ005687 was purchased from Apexbio Co. ltd (Boston, MA, USA) and dissolved in dimethyl sulfoxide (DMSO, Sigma-aldrich).

In vivo acute UTI mouse models were recapitulated by inoculating a human UPEC strain UTI89 (50 μL of 10⁸ colony forming units (CFU) in PBS) directly into the bladder of C57Bl/6J mice (female, 7-8 week-old).⁶ (link) UTI89 strain was generously provided by Dr. Scott Hultgren (Washington University, St. Louis, USA). Non-antibiotic therapeutic treatment, EPZ005687 (2.5 mg/kg, APExBIO, A4171) and DZNep (1.5 mg/kg, Selleckchem, S7120), were administered via intraperitoneal injection to wild type mice modeled for chronic cystitis and C⁺UTI superinfection; EPZ005687 is a Ezh2-specific inhibitor and DZNep is a non-specific Ezh2 inhibitor.