Exp Mol Med.  2004 Feb;36(1):43-51.

Transduction of yeast cytosine deaminase mediated by HIV-1 Tat basic domain into tumor cells induces chemosensitivity to 5-fluorocytosine

• Lee H
• Ryu J
• Kim KA
• Lee KS
• Lee JY
• Park JB
• Park J
• Choi SY

Abstract

Enzyme/prodrug approach is one of the actively developing areas for cancer therapy. In an effort to develop more effective enzyme/prodrug systems, cell-permeable cytosine deaminase was produced by fusing yeast cytosine deaminase (yCD) in frame with RKKRRQRRR domain of HIV-1 Tat which is an efficient delivery peptide of the foreign proteins into cells. The purified Tat-yCD fusion protein expressed in Escherichia coli was readily transduced into mammalian cells in a time- and dose-dependent manner. A significant level of the transduced Tat-yCD protein was recovered in the cell and was stable for 24 h as indicated by both results of the enzymatic assay of 5-fluorocytosine (5-FC) conversion to 5-fluorouracil (5-FU) and Western blot analysis. The cells transduced with Tat-yCD become highly sensitive to the cytotoxicity of 5-FC, while cells treated with yCD are unaffected by 5-FC. In addition, a strong bystander effect was observed with conditioned media from cells transduced with Tat-yCD added to non-transduced cells. Tat-yCD fusion protein demonstrated here for its ability to transduce into cells and convert nontoxic prodrug 5-FC to the toxic antimetabolite 5-FU, may be a useful approach for cancer therapy.

Keyword

cancer; cytosine deaminase; prodrug; transduction; Tat

MeSH Terms

Animals
Antimetabolites/*metabolism/pharmacology
Bystander Effect
Cytosine Deaminase/genetics/*metabolism
Flucytosine/*metabolism/pharmacology
Gene Products, tat/chemistry/genetics/*metabolism
Genetic Vectors/genetics/metabolism
HIV-1/metabolism
Hela Cells/drug effects/physiology
Humans
Prodrugs/metabolism/therapeutic use
Recombinant Fusion Proteins/genetics/*metabolism
Research Support, Non-U.S. Gov't
Saccharomyces cerevisiae Proteins/genetics/*metabolism
*Transduction, Genetic