A number of imaging marker genes have been developed for the much required need of detecting and quantitating gene expression in vivo. The overall goal of this exploratory research project is to investigate tyrosinase as new bifunctional therapeutic prodrug/imaging enzymatic marker system. The main reasons for choosing tyrosinase are 1) the potential for imaging tyrosinase expression by different imaging modalities including MR imaging(e.g. tyraminyl-DOTA-Gd) and nuclear imaging (e.g. labeled tyrosine), 2) the availability of several non-cytotoxic prodrugs and 3) the lack of expression in non-melanotic cells. We hypothesized that certain tyrosinase mutants could be obtained that would have low intrinsic toxicity, similar or even higher specific enzyme activity compared to the wild-type enzyme or could be positioned on the cell surface or be secreted for more efficient interaction with the prodrug. So far we have constructed three C-terminal tyrosinase deletion mutants lacking different elements of transmembrane and sorting domains. The novel tyrosinase variants indeed had higher specific enzyme activity compared to the wild type and sensitized transfected cells to hydroxyphenyl-propanol and N-acetyl-4-S-cysteaminylphenol treatments, i.e. fwo of several available model therapeutic prodrugs. We have furthermore confirmed that tyrosinase expressing cells accumulate/convert 3H-tyrosine opening the possibility of nuclear imaging using alternative isotopes (e.g. aboutC-tyrosine). Finally, we have synthesized a novel paramagnetic tyrosinase substrate (tyrarninyl-DOTA-Gd) which significantly changes R1 relaxivity after tyrosinase interaction. The proposed studies build on these preliminary data will test the main hypothesis that engineered tyrosinase mutants are subject to differential intracellular sorting and can be utilized both as efficient prodrugconverting biocatalysts and imaging markers. The long-term goal of this research is to explore and develop novel and useful bifunctional """"""""imaging/therapeutic marker genes"""""""" for in vivo use.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Exploratory/Developmental Grants (R21)
Project #
1R21CA085657-01A1
Application #
6266275
Study Section
Diagnostic Radiology Study Section (RNM)
Program Officer
Menkens, Anne E
Project Start
2001-02-01
Project End
2003-01-31
Budget Start
2001-02-01
Budget End
2002-01-31
Support Year
1
Fiscal Year
2001
Total Cost
$173,000
Indirect Cost
Name
Massachusetts General Hospital
Department
Type
DUNS #
City
Boston
State
MA
Country
United States
Zip Code
02199
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Simonova, Maria; Shtanko, Olena; Sergeyev, Nikolai et al. (2003) Engineering of technetium-99m-binding artificial receptors for imaging gene expression. J Gene Med 5:1056-66
Kim, Young Ro; Savellano, Mark D; Weissleder, Ralph et al. (2002) Steady-state and dynamic contrast MR imaging of human prostate cancer xenograft tumors: a comparative study. Technol Cancer Res Treat 1:489-95