Prodrug converting enzyme gene delivery by L. monocytogenes.
BACKGROUND
Listeria monocytogenes is a highly versatile bacterial carrier system for introducing protein, DNA and RNA into mammalian cells. The delivery of tumor antigens with the help of this carrier into tumor-bearing animals has been successfully carried out previously and it was recently reported that L. monocytogenes is able to colonize and replicate within solid tumors after local or even systemic injection.


METHODS
Here we report on the delivery of two prodrug converting enzymes, purine-deoxynucleoside phosphorylase (PNP) and a fusion protein consisting of yeast cytosine deaminase and uracil phosphoribosyl transferase (FCU1) into cancer cells in culture by L. monocytogenes. Transfer of the prodrug converting enzymes was achieved by bacterium mediated transfer of eukaryotic expression plasmids or by secretion of the proteins directly into the host cell cytosol by the infecting bacteria.


RESULTS
The results indicate that conversion of appropriate prodrugs to toxic drugs in the cancer cells occured after both procedures although L. monocytogenes-mediated bactofection proved to be more efficient than enzyme secretion 4T1, B16 and COS-1 tumor cells. Exchanging the constitutively PCMV-promoter with the melanoma specific P4xTETP-promoter resulted in melanoma cell-specific expression of the prodrug converting enzymes but reduced the efficiencies.


CONCLUSIONS
These experiments open the way for bacterium mediated tumor specific activation of prodrugs in live animals with tumors.
Stritzker
Jochen
J
Biocenter (Microbiology), University of Würzburg, Am Hubland, D-97074 Würzburg, Germany. jochen.stritzker@biozentrum.uni-wuerzburg.de
Pilgrim
Sabine
S
Szalay
Aladar A
AA
Goebel
Werner
W
D000818:Animals; D043525:Cytosine Deaminase / Q000378:metabolism*; D016503:Drug Delivery Systems; D004798:Enzymes / Q000378:metabolism; D015316:Genetic Therapy; D005822:Genetic Vectors; D008089:Listeria monocytogenes / Q000235:genetics* / Q000378:metabolism; D009369:Neoplasms / Q000628:therapy*; D010430:Pentosyltransferases / Q000378:metabolism*; D010957:Plasmids; D011355:Prodrugs / Q000378:metabolism*; D011683:Purine-Nucleoside Phosphorylase / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014407:Tumor Cells, Cultured
D004798:Enzymes; D011355:Prodrugs; D011993:Recombinant Fusion Proteins; D010430:Pentosyltransferases; D011683:Purine-Nucleoside Phosphorylase; C023269:uracil phosphoribosyltransferase; D043525:Cytosine Deaminase