Pharmaceutical Research

, Volume 21, Issue 5, pp 736–741

Sp1-Dependent Regulation of the RTP801 Promoter and Its Application to Hypoxia-Inducible VEGF Plasmid for Ischemic Disease

  • Minhyung Lee
  • Malavosklish Bikram
  • Seungjoon Oh
  • David A. Bull
  • Sung Wan Kim
Article

DOI: 10.1023/B:PHAM.0000026421.09367.b3

Cite this article as:
Lee, M., Bikram, M., Oh, S. et al. Pharm Res (2004) 21: 736. doi:10.1023/B:PHAM.0000026421.09367.b3

Abstract

Purpose. Gene therapy using vascular endothelial growth factor (VEGF) is a new potential treatment of ischemic disease. To be safe and effective, VEGF expression should be enhanced locally in ischemic tissue. In this study, we identified the cis-regulatory element for the hypoxia induction of the RTP801 promoter. In addition, pRTP801-VEGF was evaluated as a therapeutic plasmid in vitro.

Methods. The cis-regulatory element for hypoxia induction was identified by deletion and mutation analyses. Antisense oligonucleotide co-transfection assay was performed to evaluate the role of Sp1. pRTP801-VEGF was constructed by the insertion of the RTP801 promoter into the VEGF plasmid. The hypoxia-inducible expression of VEGF was evaluated by in vitro transfection assay.

Results. In luciferase assay, the region between -495 and -446 was responsible for the hypoxia-induced transcription. The mutation of the Sp1 site in this region reduced hypoxia-induced transcription. In addition, co-transfection with antisense Sp1 oligonucleotide suggests that hypoxia induction of the RTP801 promoter is mediated by Sp1. In vitro transfection showed that pRTP801-VEGF had higher VEGF expression than pEpo-SV-VEGF. In addition, VEGF expression by pRTP801-VEGF was induced under hypoxia.

Conclusions. With strong basal promoter activity and induction under hypoxia, pRTP801-VEGF may be useful for gene therapy for ischemic disease.

hypoxiaRTP801Sp1transcriptional regulationvascular endothelial growth factor

Copyright information

© Plenum Publishing Corporation 2004

Authors and Affiliations

  • Minhyung Lee
    • 1
  • Malavosklish Bikram
    • 2
  • Seungjoon Oh
    • 3
  • David A. Bull
    • 4
  • Sung Wan Kim
    • 2
  1. 1.Clinical Research Center, College of MedicineInha UniversityInchonKorea
  2. 2.Center for Controlled Chemical Delivery, Department of Pharmaceutics and Pharmaceutical ChemistryUniversity of UtahSalt Lake CityUSA
  3. 3.Department of Internal Medicine, College of MedicineKyung Hee UniversitySeoulKorea
  4. 4.Department of Surgery, Division of Cardiothoracic SurgeryUniversity of Utah Health Sciences CenterSalt Lake CityUSA