In vivo antitumor activity of 5-fluorocytosine on human colorectal carcinoma cells genetically modified to express cytosine deaminase.
A human colorectal carcinoma cell line, WiDr, was genetically engineered to express the nonmammalian enzyme, cytosine deaminase (CD). Expression of CD in WiDr cells (WiDr/CD) did not alter the growth rate of these cells when grown in vitro or as solid tumor xenografts in nude mice. However, expression of CD did increase the sensitivity of these cells to the nontoxic prodrug, 5-fluorocytosine (FCyt), decreasing the 50% inhibitory concentration for FCyt from 26,000 microM in parental WiDr cells to 27 microM in WiDr/CD cells. The increase in sensitivity to FCyt in WiDr/CD cells was the result of the CD-mediated conversion of FCyt to 5-fluorouracil (FUra) and subsequent FUra anabolites. The half-life of the prodrug, FCyt, was determined to be approximately 40 min in nude mice. A single i.p. injection of 500 mg FCyt/kg body weight resulted in a transient FCyt plasma level of approximately 4000 microM while osmotic minipumps or constant tail vein infusions of FCyt achieved continual FCyt plasma levels of 5 microM and 50 microM, respectively, with no overt signs of toxicity. Significant antitumor effects were observed in nude mice bearing tumors derived from WiDr/CD cells when these animals were given 500 mg FCyt/kg i.p. for 10 consecutive days. These antitumor effects were demonstrated by decreases in tumor growth rate, tumor size, tumor weight, and thymidine incorporation into tumor DNA. This antitumor effect was significant but less profound if FCyt was administered by constant tail vein infusion. WiDr and WiDr/CD cells were very sensitive to FUra in vitro (50% inhibitory concentration approximately 5 microM). However, no significant antitumor effects were observed in nude mice bearing tumors derived from either WiDr or WiDr/CD cells when these animals were treated with various doses of FUra. Taken collectively, these data indicate that nontoxic plasma levels of FCyt can be attained which can produce profound antitumor effects on tumors engineered to express CD and that these antitumor effects are significantly better than those that can be achieved using FUra. These positive data support the continued development of a gene therapy approach to colorectal carcinoma involving the selective expression of CD in colorectal tumors with subsequent administration of FCyt.
Huber
B E
BE
Division of Cell Biology, Wellcome Research Laboratories, Research Triangle Park, North Carolina 27709.
Austin
E A
EA
Good
S S
SS
Knick
V C
VC
Tibbels
S
S
Richards
C A
CA
D000818:Animals; D002455:Cell Division / Q000187:drug effects; D015179:Colorectal Neoplasms / Q000188:drug therapy* / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D043525:Cytosine Deaminase; D005260:Female; D005437:Flucytosine / Q000493:pharmacokinetics / Q000627:therapeutic use* / Q000633:toxicity; D005472:Fluorouracil / Q000493:pharmacokinetics / Q000627:therapeutic use* / Q000633:toxicity; D006801:Humans; D007700:Kinetics; D051379:Mice; D008819:Mice, Nude; D009368:Neoplasm Transplantation; D009700:Nucleoside Deaminases / Q000096:biosynthesis* / Q000235:genetics; D014162:Transfection; D014183:Transplantation, Heterologous; D014407:Tumor Cells, Cultured
D005437:Flucytosine; D009700:Nucleoside Deaminases; D043525:Cytosine Deaminase; D005472:Fluorouracil