Molecular Imaging and Biology

, Volume 16, Issue 2, pp 224–234

A Titratable Two-Step Transcriptional Amplification Strategy for Targeted Gene Therapy Based on Ligand-Induced Intramolecular Folding of a Mutant Human Estrogen Receptor

  • Ian Y. Chen
  • Ramasamy Paulmurugan
  • Carsten H. Nielsen
  • David S. Wang
  • Vinca Chow
  • Robert C. Robbins
  • Sanjiv S. Gambhir
Research Article

Abstract

Purpose

The efficacy and safety of cardiac gene therapy depend critically on the level and the distribution of therapeutic gene expression following vector administration. We aimed to develop a titratable two-step transcriptional amplification (tTSTA) vector strategy, which allows modulation of transcriptionally targeted gene expression in the myocardium.

Procedures

We constructed a tTSTA plasmid vector (pcTnT-tTSTA-fluc), which uses the cardiac troponin T (cTnT) promoter to drive the expression of the recombinant transcriptional activator GAL4-mER(LBD)-VP2, whose ability to transactivate the downstream firefly luciferase reporter gene (fluc) depends on the binding of its mutant estrogen receptor (ERG521T) ligand binding domain (LBD) to an ER ligand such as raloxifene. Mice underwent either intramyocardial or hydrodynamic tail vein (HTV) injection of pcTnT-tTSTA-fluc, followed by differential modulation of fluc expression with varying doses of intraperitoneal raloxifene prior to bioluminescence imaging to assess the kinetics of myocardial or hepatic fluc expression.

Results

Intramyocardial injection of pcTnT-tTSTA-fluc followed by titration with intraperitoneal raloxifene led to up to tenfold induction of myocardial fluc expression. HTV injection of pcTnT-tTSTA-fluc led to negligible long-term hepatic fluc expression, regardless of the raloxifene dose given.

Conclusions

The tTSTA vector strategy can effectively modulate transgene expression in a tissue-specific manner. Further refinement of this strategy should help maximize the benefit-to-risk ratio of cardiac gene therapy.

Key words

Gene therapy Drug-regulated gene expression Transcriptional amplification Transcriptional targeting Intramolecular folding Bioluminescence imaging 

References

  1. 1.
    Mitsos S, Katsanos K, Koletsis E et al (2012) Therapeutic angiogenesis for myocardial ischemia revisited: basic biological concepts and focus on latest clinical trials. Angiogenesis 15:1–22PubMedCrossRefGoogle Scholar
  2. 2.
    Franz WM, Breves D, Klingel K et al (1993) Heart-specific targeting of firefly luciferase by the myosin light chain-2 promoter and developmental regulation in transgenic mice. Circ Res 73:629–638PubMedCrossRefGoogle Scholar
  3. 3.
    Chen IY, Gheysens O, Ray S et al (2010) Indirect imaging of cardiac-specific transgene expression using a bidirectional two-step transcriptional amplification strategy. Gene Ther 17:827–838PubMedCentralPubMedCrossRefGoogle Scholar
  4. 4.
    Paulmurugan R, Gambhir SS (2006) An intramolecular folding sensor for imaging estrogen receptor-ligand interactions. Proc Natl Acad Sci U S A 103:15883–15888PubMedCentralPubMedCrossRefGoogle Scholar
  5. 5.
    von Degenfeld G, Banfi A, Springer ML et al (2006) Microenvironmental VEGF distribution is critical for stable and functional vessel growth in ischemia. FASEB J 20:2657–2659CrossRefGoogle Scholar
  6. 6.
    Ozawa CR, Banfi A, Glazer NL et al (2004) Microenvironmental VEGF concentration, not total dose, determines a threshold between normal and aberrant angiogenesis. J Clin Invest 113:516–527PubMedCentralPubMedCrossRefGoogle Scholar
  7. 7.
    Yla-Herttuala S, Rissanen TT, Vajanto I, Hartikainen J (2007) Vascular endothelial growth factors: biology and current status of clinical applications in cardiovascular medicine. J Am Coll Cardiol 49:1015–1026PubMedCrossRefGoogle Scholar
  8. 8.
    Reiss K, Cheng W, Ferber A et al (1996) Overexpression of insulin-like growth factor-1 in the heart is coupled with myocyte proliferation in transgenic mice. Proc Natl Acad Sci U S A 93:8630–8635PubMedCentralPubMedCrossRefGoogle Scholar
  9. 9.
    Nuss HB, Marban E, Johns DC (1999) Overexpression of a human potassium channel suppresses cardiac hyperexcitability in rabbit ventricular myocytes. J Clin Invest 103:889–896PubMedCentralPubMedCrossRefGoogle Scholar
  10. 10.
    Bish LT, Sleeper MM, Reynolds C et al (2011) Cardiac gene transfer of short hairpin RNA directed against phospholamban effectively knocks down gene expression but causes cellular toxicity in canines. Hum Gene Ther 22:969–977PubMedCentralPubMedCrossRefGoogle Scholar
  11. 11.
    Goldstein SR, Siddhanti S, Ciaccia AV, Plouffe L Jr (2000) A pharmacological review of selective oestrogen receptor modulators. Hum Reprod Update 6:212–224PubMedCrossRefGoogle Scholar
  12. 12.
    Kocanova S, Mazaheri M, Caze-Subra S, Bystricky K (2010) Ligands specify estrogen receptor alpha nuclear localization and degradation. BMC Cell Biol 11:98PubMedCentralPubMedCrossRefGoogle Scholar
  13. 13.
    Xiong W, Goverdhana S, Sciascia SA et al (2006) Regulatable gutless adenovirus vectors sustain inducible transgene expression in the brain in the presence of an immune response against adenoviruses. J Virol 80:27–37PubMedCentralPubMedCrossRefGoogle Scholar
  14. 14.
    Fleury S, Driscoll R, Simeoni E et al (2004) Helper-dependent adenovirus vectors devoid of all viral genes cause less myocardial inflammation compared with first-generation adenovirus vectors. Basic Res Cardiol 99:247–256PubMedGoogle Scholar
  15. 15.
    Pleger ST, Shan C, Ksienzyk J et al (2011) Cardiac AAV9-S100A1 gene therapy rescues post-ischemic heart failure in a preclinical large animal model. Sci Transl Med 3:92ra64PubMedCrossRefGoogle Scholar
  16. 16.
    Rodriguez-Porcel M, Brinton TJ, Chen IY et al (2008) Reporter gene imaging following percutaneous delivery in swine moving toward clinical applications. J Am Coll Cardiol 51:595–597PubMedCentralPubMedCrossRefGoogle Scholar
  17. 17.
    Lee SW, Lee SH, Biswal S (2012) Magnetic resonance reporter gene imaging. Theranostics 2:403–412PubMedCentralPubMedCrossRefGoogle Scholar
  18. 18.
    Herweijer H, Zhang G, Subbotin VM et al (2001) Time course of gene expression after plasmid DNA gene transfer to the liver. J Gene Med 3:280–291PubMedCrossRefGoogle Scholar
  19. 19.
    Son MK, Choi JH, Lee DS et al (2005) Pharmacokinetics and biodistribution of a pGT2-VEGF plasmid DNA after administration in rats. J Cardiovasc Pharmacol 46:577–584PubMedCrossRefGoogle Scholar
  20. 20.
    Heindel JJ, vom Saal FS (2008) Meeting report: batch-to-batch variability in estrogenic activity in commercial animal diets—importance and approaches for laboratory animal research. Environ Health Perspect 116:389–393PubMedCentralPubMedCrossRefGoogle Scholar
  21. 21.
    Yang Z, He X, Zhang Y (2007) The determination of raloxifene in rat tissue using HPLC. Biomed Chrom BMC 21:229–233CrossRefGoogle Scholar
  22. 22.
    Kuiper GG, Carlsson B, Grandien K et al (1997) Comparison of the ligand binding specificity and transcript tissue distribution of estrogen receptors alpha and beta. Endocrinology 138:863–870PubMedGoogle Scholar
  23. 23.
    Guo ZS, Li Q, Bartlett DL, Yang JY, Fang B (2008) Gene transfer: the challenge of regulated gene expression. Trends Mol Med 14:410–418PubMedCrossRefGoogle Scholar
  24. 24.
    Muller OJ, Katus HA, Bekeredjian R (2007) Targeting the heart with gene therapy-optimized gene delivery methods. Cardiovasc Res 73:453–462PubMedCrossRefGoogle Scholar

Copyright information

© World Molecular Imaging Society 2013

Authors and Affiliations

  • Ian Y. Chen
    • 1
  • Ramasamy Paulmurugan
    • 2
  • Carsten H. Nielsen
    • 3
  • David S. Wang
    • 2
  • Vinca Chow
    • 4
  • Robert C. Robbins
    • 5
  • Sanjiv S. Gambhir
    • 2
    • 6
    • 7
    • 8
  1. 1.Department of Medicine, Division of Cardiovascular Medicine, Stanford University School of MedicineStanford UniversityStanfordUSA
  2. 2.Department of Radiology, Molecular Imaging Program at Stanford (MIPS), Stanford University School of MedicineStanford UniversityStanfordUSA
  3. 3.Cluster for Molecular Imaging, Department of Clinical Physiology, Nuclear Medicine and PET, RigshospitaletUniversity of CopenhagenCopenhagenDenmark
  4. 4.Department of AnesthesiologyBrigham and Women’s HospitalBostonUSA
  5. 5.Department of Cardiothoracic Surgery, Stanford University School of MedicineStanford UniversityStanfordUSA
  6. 6.The Bio-X Program, Stanford University School of MedicineStanford UniversityStanfordUSA
  7. 7.Department of BioengineeringStanford UniversityStanfordUSA
  8. 8.Department of Material Science and EngineeringStanford UniversityStanfordUSA

Personalised recommendations