Preferential cytotoxicity of cells transduced with cytosine deaminase compared to bystander cells after treatment with 5-flucytosine.
In vitro experiments from our laboratory and others have suggested that herpes simplex virus thymidine kinase (HSV-TK)/ganciclovir (GCV) gene therapy depends on gap junctional intercellular communication (GJIC) to produce a strong bystander effect. Furthermore, we have shown that cells transduced with HSV-TK can be protected from GCV-mediated toxicity by GJIC with bystander cells. We wished to determine whether GJIC affected either the bystander or protective effect of the cytosine deaminase (CD)/5-flucytosine (5-FC) gene therapy approach, in which CD converts 5-FC to 5-fluorouracil (5-FU). To test this, we designed a coculture system using communication-competent WB rat hepatocytes and a noncommunicating subclone (aB1), which were transduced with CD and with antibiotic resistance genes so that we could independently determine the survival of the CD-containing or bystander cells. We found that, compared to the HSV-TK/GCV strategy, bystander killing resulting from treatment with CD/5-FC does not depend on GJIC. However, our most striking finding was that both communication-competent and -incompetent CD-transduced cells were preferentially killed, by a factor of up to 500, compared to bystander cells. The lesser dependence of the CD/5-FC system on GJIC, combined with the finding that most cancer cells lack the capacity for GJIC, suggest that the CD/5-FC system may be superior to the HSV-TK/GCV approach for gene therapy. However, the premature death of the CD-transduced 5-FU "factory" suggests that other strategies may be necessary to produce a sufficient quantity of 5-FU for a duration long enough to produce permanent tumor regression.
Lawrence
T S
TS
Department of Radiation Oncology, University of Michigan Medical Center, Ann Arbor 48109-0010, USA. tsl@umich.edu
Rehemtulla
A
A
Ng
E Y
EY
Wilson
M
M
Trosko
J E
JE
Stetson
P L
PL
D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000378:metabolism / Q000494:pharmacology*; D002470:Cell Survival / Q000187:drug effects*; D043525:Cytosine Deaminase; D005437:Flucytosine / Q000378:metabolism / Q000494:pharmacology*; D017629:Gap Junctions; D015316:Genetic Therapy; D006801:Humans; D009700:Nucleoside Deaminases / Q000235:genetics / Q000494:pharmacology*; D011355:Prodrugs / Q000378:metabolism / Q000494:pharmacology*; D051381:Rats; D014407:Tumor Cells, Cultured / Q000187:drug effects
D000964:Antimetabolites, Antineoplastic; D011355:Prodrugs; D005437:Flucytosine; D009700:Nucleoside Deaminases; D043525:Cytosine Deaminase