1. ¹The Crump Institute for Molecular Imaging, David Geffen School of Medicine, University of California at Los Angeles, Los Angeles, CA 90095, USA
2. ²Department of Molecular and Medical Pharmacology, David Geffen School of Medicine, University of California at Los Angeles, Los Angeles, CA 90095, USA
3. ³Department of Biological Chemistry, David Geffen School of Medicine, University of California at Los Angeles, Los Angeles, CA 90095, USA
4. ⁴Department of Urology, David Geffen School of Medicine, University of California at Los Angeles, Los Angeles, CA 90095, USA
5. ⁵Department of Radiology and Bio-X Program, Molecular Imaging Program at Stanford, Stanford University School of Medicine, Stanford, CA 94305, USA

Correspondence: Sanjiv S. Gambhir, James H. Clark Center, Stanford University School of Medicine, 318 Campus Drive, 1E, Stanford, CA 94305-5427, USA. E-mail: sgambhir@stanford.edu

Received 9 February 2004; Accepted 5 June 2004.

Abstract

Noninvasive evaluation of gene transfer to specific cells or tissues will allow for long-term, repetitive monitoring of transgene expression. Tissue-specific promoters that restrict the expression of a transgene to tumor cells play a vital role in cancer gene therapy imaging. In this study, we have developed a third-generation HIV-1-based lentivirus vector carrying a prostate-specific promoter to monitor the long-term, sustained expression of the firefly luciferase (fl) reporter gene in living mice. The fl gene in the transcriptionally targeted vector is driven by an enhanced prostate-specific antigen promoter in a two-step transcriptional amplification (TSTA) system. The efficiency of the lentivirus (LV-TSTA)-mediated gene delivery, cell-type specificity, and persistence of gene expression were evaluated in cell culture and in living mice carrying prostate tumor xenografts. In vivo bioluminescence imaging with a cooled charge-coupled device camera revealed significantly high levels of fl expression in prostate tumors. Injection of LV-TSTA directly into the prostate of male nude mice revealed efficient and long-term fl gene expression in the prostate tissue for up to 3 months. These studies demonstrate the significant potential of TSTA-based lentivirus vectors to confer high levels of tissue-specific gene expression from a weak promoter, while preserving cell-type specificity and the ability to image noninvasively the sustained, long-term expression of reporter genes in living animals.