Concomitant expression of E. coli cytosine deaminase and uracil phosphoribosyltransferase improves the cytotoxicity of 5-fluorocytosine.
The prodrug activation system formed by the E. coli codA gene encoding cytosine deaminase (CD) and 5-fluorocytosine (5-FC) developed for selective cancer chemotherapy suffers from a sensitivity limitation in many tumour cells. In an attempt to improve the CD/5-FC suicide association, we combined the E. coli upp gene encoding uracil phosphoribosyltransferase (UPRT) with codA gene to create the situation prevailing in E. coli, a bacterium very efficient in metabolising 5-FC. The constitutive expression of the two genes cloned on an E. coli-animal cell shuttle plasmid either in a linked or in a fused configuration was evaluated in E. coli strains selected and engineered to mimic the 5-FC metabolism encountered in mammalian cells. The simultaneous expression of codA and upp genes generated a cooperative effect resulting in a dramatic increase in 5-FC sensitivity of cells compared to the expression of codA alone. Furthermore, it was shown that the association of UPRT with CD facilitated the uptake of 5-FC, in the situation where the drug penetrates cells by passive diffusion as in mammalian cells, by directly channeling 5-fluorouracil, the product of CD, to 5-fluoroUMP, the product of UPRT.
Tiraby
M
M
CAYLA, Toulouse, France. tiraby@cict.fr
Cazaux
C
C
Baron
M
M
Drocourt
D
D
Reynes
J P
JP
Tiraby
G
G
D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000378:metabolism / Q000494:pharmacology; D019677:Artificial Gene Fusion; D043525:Cytosine Deaminase; D004351:Drug Resistance / Q000235:genetics; D004926:Escherichia coli / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics*; D029968:Escherichia coli Proteins; D005437:Flucytosine / Q000378:metabolism* / Q000494:pharmacology*; D005472:Fluorouracil / Q000494:pharmacology; D015870:Gene Expression; D005798:Genes, Bacterial; D009154:Mutation; D009374:Neoplasms, Experimental / Q000188:drug therapy / Q000378:metabolism; D009700:Nucleoside Deaminases / Q000235:genetics* / Q000378:metabolism; D010430:Pentosyltransferases / Q000235:genetics* / Q000378:metabolism; D011355:Prodrugs / Q000378:metabolism
D000964:Antimetabolites, Antineoplastic; D029968:Escherichia coli Proteins; D011355:Prodrugs; D005437:Flucytosine; D010430:Pentosyltransferases; C023269:uracil phosphoribosyltransferase; D009700:Nucleoside Deaminases; D043525:Cytosine Deaminase; C468827:codA protein, E coli; D005472:Fluorouracil