Suppression of VEGF transcription in renal cell carcinoma cells by pyrrole-imidazole hairpin polyamides targeting the hypoxia responsive element.
Hypoxia inducible factor (HIF), a master regulator of critical genes for cell survival under hypoxic conditions, is known to be related to tumorigenesis and progression of renal cell carcinoma. N-methylpyrrole (Py)-N-methylimidazole (Im) hairpin polyamides are synthetic organic compounds that recognize and bind to the minor grooves of specific DNA sequences. We synthesized three Py-Im hairpin polyamides targeting the flanking sequences of hypoxia responsive element (HRE; a binding site of HIF) in the promoter region of the vascular endothelial growth factor (VEGF) gene. The effects of the polyamides on HIF-induced transcription were evaluated by a luciferase assay using a reporter plasmid containing a VEGF promoter. Real time reverse-transcriptase polymerase chain reaction and enzyme-linked immunosorbent assay were performed to examine the effects of the polyamides on the transcription and secretion of VEGF in A498 renal cell carcinoma cells, which have a frame-shift mutation in the von Hippel-Lindau gene. A combination of three Py-Im hairpin polyamides suppressed HIF-induced transcription in reporter assays using 293 cells and successfully suppressed transcription and translation of the VEGF gene in A498 cells. Inhibition of the HIF-HRE interaction was confirmed by an electrophoresis mobility shift assay. An approach using Py-Im hairpin polyamides may be a new strategy for the treatment of renal cell carcinoma.
Kageyama
Yukio
Y
Department of Urology, Graduate School of Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo, 113-8519, Japan. kageyys.uro@tmd.ac.jp
Sugiyama
Hiroshi
H
Ayame
Hirohito
H
Iwai
Aki
A
Fujii
Yasuhisa
Y
Huang
L Eric
LE
Kizaka-Kondoh
Shinae
S
Hiraoka
Masahiro
M
Kihara
Kazunori
K
D001483:Base Sequence; D051792:Basic Helix-Loop-Helix Proteins; D002292:Carcinoma, Renal Cell / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism*; D015687:Cell Hypoxia / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D004359:Drug Therapy, Combination; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism; D007093:Imidazoles / Q000737:chemistry; D008954:Models, Biological; D008969:Molecular Sequence Data; D009757:Nylons / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D011758:Pyrroles / Q000737:chemistry; D020218:Response Elements / Q000187:drug effects*; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D014407:Tumor Cells, Cultured; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics* / Q000378:metabolism*; D000099162:Endothelial PAS Domain-Containing Protein 1
D051792:Basic Helix-Loop-Helix Proteins; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007093:Imidazoles; D009757:Nylons; D011758:Pyrroles; D014157:Transcription Factors; D042461:Vascular Endothelial Growth Factor A; D000099162:Endothelial PAS Domain-Containing Protein 1; C497442:HIF1A protein, human; C467484:VEGFA protein, human; C029899:imidazole