A novel gene silencer, pyrrole-imidazole polyamide targeting human lectin-like oxidized low-density lipoprotein receptor-1 gene improves endothelial cell function.
Pyrrole-imidazole polyamide can be combined in antiparallel side-by-side dimeric complexes along the minor groove of DNA in a sequence-specific manner. Pyrrole-imidazole polyamides are effective inhibitors of transcription factors as well as viral repressors and transactivators. Recently, lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) was reported to be a major factor contributing to the pathogenesis of coronary atherosclerosis. In this study, we designed a pyrrole-imidazole polyamide specific for the LOX-1 gene and evaluated its effect on LOX-1 gene transcription. A pyrrole-imidazole polyamide was designed to target the AP-1 binding site of the LOX-1 gene and synthesized by solid phase methods. This pyrrole-imidazole polyamide significantly inhibited LOX-1 promoter activity in HEK293 cells, determined by the luciferase assay. LOX-1 mRNA expression was also inhibited by the pyrrole-imidazole polyamide at a concentration of 10-9 mol/l in human umbilical vein endothelial cells (HUVEC), determined by the real-time PCR method. HUVEC were treated by pyrrole-imidazole polyamide targeting the LOX-1 gene, and apoptosis was assessed using Hoechst stain, terminal deoxy nucleotidyl transferase-mediated UTP end labeling method, and dye-uptake bioassay. Treatment of HUVEC for 72 h with LOX-1 targeted pyrrole-imidazole polyamide decreased apoptosis induced by angiotensin II and oxidized low-density lipoprotein (ox-LDL) loading in all assays. This novel therapeutic agent, pyrrole-imidazole polyamide, could specifically inhibit LOX-1 gene expression by reducing the promoter activity of the gene. Pyrrole-imidazole polyamide seems to be a powerful promising new agent that can be used to explore therapies based on inhibition of transcription. Molecular recognition of DNA by small molecules could provide insight into the development of new human medicines.
Ueno
Takahiro
T
Department of Medicine, Division of Nephrology, Hypertension and Endocrinology, Itabashi, Nihon University School of Medicine, Itabashi, Tokyo, Japan. tueno@med.nihon-u.ac.jp
Fukuda
Noboru
N
Tsunemi
Akiko
A
Yao
En-Hui
EH
Matsuda
Hiroyuki
H
Tahira
Kazunobu
K
Matsumoto
Taro
T
Matsumoto
Koichi
K
Matsumoto
Yoshiaki
Y
Nagase
Hiroki
H
Sugiyama
Hiroshi
H
Sawamura
Tatsuya
T