Novel bidirectional vector strategy for amplification of therapeutic and reporter gene expression

Human Gene Therapy
S RaySanjiv S Gambhir

Abstract

Molecular imaging methods have previously been employed to image tissue-specific reporter gene expression by a two-step transcriptional amplification (TSTA) strategy. We have now developed a new bidirectional vector system, based on the TSTA strategy, that can simultaneously amplify expression for both a target gene and a reporter gene, using a relatively weak promoter. We used the synthetic Renilla luciferase (hrl) and firefly luciferase (fl) reporter genes to validate the system in cell cultures and in living mice. When mammalian cells were transiently cotransfected with the GAL4-responsive bidirectional reporter vector and various doses of the activator plasmid encoding the GAL4-VP16 fusion protein, pSV40-GAL4-VP16, a high correlation (r(2) = 0.95) was observed between the expression levels of both reporter genes. Good correlations (r(2) = 0.82 and 0.66, respectively) were also observed in vivo when the transiently transfected cells were implanted subcutaneously in mice or when the two plasmids were delivered by hydrodynamic injection and imaged. This work establishes a novel bidirectional vector approach utilizing the TSTA strategy for both target and reporter gene amplification. This validated approach should prove useful ...Continue Reading

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Citations

Jan 11, 2005·The Journal of Pathology·Scott K Lyons
Sep 24, 2005·European Journal of Nuclear Medicine and Molecular Imaging·Iván PeñuelasSanjiv S Gambhir
Jun 23, 2006·Molecular Imaging and Biology : MIB : the Official Publication of the Academy of Molecular Imaging·Meike L SchipperSanjiv S Gambhir
Jun 26, 2008·Molecular Therapy : the Journal of the American Society of Gene Therapy·Ekaterina V FomichevaJoseph M Metzger
Sep 4, 2008·Molecular Therapy : the Journal of the American Society of Gene Therapy·Sunetra RaySanjiv S Gambhir
Aug 6, 2009·Molecular Therapy : the Journal of the American Society of Gene Therapy·Ramasamy PaulmuruganSanjiv S Gambhir
Dec 19, 2012·Molecular Therapy : the Journal of the American Society of Gene Therapy·Frédéric PouliotLily Wu
Oct 4, 2007·Molecular Therapy : the Journal of the American Society of Gene Therapy·Takeshi Suda, Dexi Liu
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Dec 17, 2010·Cancer Research·Manishkumar R PatelSanjiv S Gambhir
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Mar 25, 2006·Future Oncology·John Y Dong, Jan Woraratanadharm
Dec 7, 2006·Neuroimaging Clinics of North America·Dawid Schellingerhout
Aug 15, 2006·Molecular Therapy : the Journal of the American Society of Gene Therapy·B H LiuS Wang
Dec 15, 2006·Gene Therapy·H Herweijer, J A Wolff
Jun 22, 2006·Journal of Orthopaedic Research : Official Publication of the Orthopaedic Research Society·Brian T FeeleyJay R Lieberman
Nov 9, 2005·Molecular Cancer Therapeutics·Romyla IlaganMichael Carey
Dec 3, 2009·Molecular Cancer Therapeutics·Maja L ArendtIain M Morgan

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