Synthetic pyrrole-imidazole polyamide inhibits expression of the human transforming growth factor-beta1 gene

• Published in: The Journal of pharmacology and experimental therapeutics Vol. 315; no. 2; pp. 571 - 575
• Main Authors: Lai, Yu-Mu, Fukuda, Noboru, Ueno, Takahiro, Matsuda, Hiroyuki, Saito, Satoshi, Matsumoto, Koichi, Ayame, Hirohito, Bando, Toshikazu, Sugiyama, Hiroshi, Mugishima, Hideo, Serie, Kazuo
• Format: Journal Article
• Language: English
• Published: United States 01.11.2005
• Subjects: Blotting, Western; Cells, Cultured; Deoxyribonucleases, Type II Site-Specific - biosynthesis; Deoxyribonucleases, Type II Site-Specific - genetics; DNA - metabolism; Electrophoretic Mobility Shift Assay; Endothelial Cells - drug effects; Endothelial Cells - metabolism; Gene Expression Regulation - drug effects; Humans; Indicators and Reagents; Nylons - chemical synthesis; Nylons - pharmacology; Oligonucleotides - metabolism; Promoter Regions, Genetic; Pyrroles - chemical synthesis; Pyrroles - pharmacology; Reverse Transcriptase Polymerase Chain Reaction; Transforming Growth Factor beta - biosynthesis; Transforming Growth Factor beta - genetics; Transforming Growth Factor beta1; Tubulin - biosynthesis; Tubulin - genetics
• ISSN: 0022-3565

Pyrrole-imidazole (Py-Im) polyamides can bind to the predetermined base pairs in the minor groove of double-helical DNA with high affinity. These synthetic small molecules can interfere with transcription factor-DNA interaction and inhibit or activate the transcription of corresponding genes. In the present study, we designed and synthesized a Py-Im polyamide to target -545 to -539 base pairs of human transforming growth factor-beta1 (hTGF-beta1) promoter adjacent to the fat-specific element 2 (FSE2) to inhibit the expression of the gene. Gel mobility shift assay showed that the synthetic Py-Im polyamide binds to its corresponding double-strand oligonucleotides, whereas the mismatch polyamides did not bind. Fluorescein isothiocyanate-labeled Py-Im polyamide was detected in the nuclei of human vascular smooth muscle cells (VSMCs) after 2- to 48-h incubation. Py-Im polyamide significantly decreased the promoter activity of hTGF-beta1 determined by in vitro transcription experiments and luciferase assay. In cultured human VSMCs, Py-Im polyamide targeting hTGF-beta1 promoter significantly inhibited expressions of hTGF-beta1 mRNA and protein. These results indicate that the synthetic Py-Im polyamide designed to bind hTGF-beta1 promoter inhibited hTGF-beta1 gene and protein expression successfully. This novel agent will be used for the TGF-beta-related diseases as a gene therapy.