The overall goal of this research proposal is to develop a widely applicable method for noninvasive imaging of gene expression in target tissue transduced by a multiple gene delivery vector. The applicants have previously demonstrated that HSV1-tk gene expression can be measured and imaged with currently available clinical equipment using radiolabeled substrates that are selectively metabolized by HSV1-tk and accumulate in transduced tissue. The current proposal seeks to demonstrate that imaging gene expression can be extended to a broad range of genes that are of therapeutic interest. The approach will use multiple gene delivery vectors that include HSV1-tk as a """"""""marker gene"""""""" and a different """"""""therapeutic gene"""""""" in the same expression cassette. The focus of this proposal is to demonstrate that proportional and consistent co-expression of both """"""""therapeutic"""""""" and """"""""marker"""""""" genes can be achieved in target tissue (over a wide range of gene expression) following transduction with a multiple gene retroviral vector. The applicants propose to test several existing multiple gene delivery vector constructs (multiple gene expression cassettes) for their ability to provide sufficient and proportional levels of co-expression of two or more genes in transduced tissue. The research will be directed at evaluating different multiple gene delivery vector constructs that include HSV1-tk (as a marker gene) and three genes of therapeutic interest [double mutant dihydrofolate reductase (DHFR), mutant low affinity nerve growth factor receptor (NTP), and granulocyte-macrophage colony stimulating factor (GM-CSF)] that have different sites of gene product activity (intracellular, cell membrane, and extracellular). By including activator-repressor elements in the bicistronic vectors, the applicants will also test the applicability of noninvasive imaging of HSV1-tk expression to monitor changes in gene expression induced by exogenous modulatory stimuli. These studies will be performed first in tissue culture and then in animals using clinical imaging techniques.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA076117-05
Application #
6513140
Study Section
Diagnostic Radiology Study Section (RNM)
Program Officer
Menkens, Anne E
Project Start
1998-07-01
Project End
2004-04-30
Budget Start
2002-05-01
Budget End
2004-04-30
Support Year
5
Fiscal Year
2002
Total Cost
$260,665
Indirect Cost
Name
Sloan-Kettering Institute for Cancer Research
Department
Type
DUNS #
064931884
City
New York
State
NY
Country
United States
Zip Code
10065
Zanzonico, Pat; Koehne, Guenther; Gallardo, Humilidad F et al. (2006) [131I]FIAU labeling of genetically transduced, tumor-reactive lymphocytes: cell-level dosimetry and dose-dependent toxicity. Eur J Nucl Med Mol Imaging 33:988-97
Serganova, Inna; Doubrovin, Michael; Vider, Jelena et al. (2004) Molecular imaging of temporal dynamics and spatial heterogeneity of hypoxia-inducible factor-1 signal transduction activity in tumors in living mice. Cancer Res 64:6101-8
Doubrovin, Mikhail; Ponomarev, Vladimir; Serganova, Inna et al. (2003) Development of a new reporter gene system--dsRed/xanthine phosphoribosyltransferase-xanthine for molecular imaging of processes behind the intact blood-brain barrier. Mol Imaging 2:93-112
Koehne, Guenther; Doubrovin, Mikhail; Doubrovina, Ekaterina et al. (2003) Serial in vivo imaging of the targeted migration of human HSV-TK-transduced antigen-specific lymphocytes. Nat Biotechnol 21:405-13
Tjuvajev, Juri Gelovani; Doubrovin, Mikhail; Akhurst, Timothy et al. (2002) Comparison of radiolabeled nucleoside probes (FIAU, FHBG, and FHPG) for PET imaging of HSV1-tk gene expression. J Nucl Med 43:1072-83
Ponomarev, V; Doubrovin, M; Lyddane, C et al. (2001) Imaging TCR-dependent NFAT-mediated T-cell activation with positron emission tomography in vivo. Neoplasia 3:480-8
Jacobs, A; Tjuvajev, J G; Dubrovin, M et al. (2001) Positron emission tomography-based imaging of transgene expression mediated by replication-conditional, oncolytic herpes simplex virus type 1 mutant vectors in vivo. Cancer Res 61:2983-95
Doubrovin, M; Ponomarev, V; Beresten, T et al. (2001) Imaging transcriptional regulation of p53-dependent genes with positron emission tomography in vivo. Proc Natl Acad Sci U S A 98:9300-5
Jacobs, A; Dubrovin, M; Hewett, J et al. (1999) Functional coexpression of HSV-1 thymidine kinase and green fluorescent protein: implications for noninvasive imaging of transgene expression. Neoplasia 1:154-61