Tumor uptake and elimination of 2',2'-difluoro-2'-deoxycytidine (gemcitabine) after deoxycytidine kinase gene transfer: correlation with in vivo tumor response.
OBJECTIVE
We hypothesized that tumor uptake and elimination of 2',2'-difluoro-2'-deoxycytidine/2',2'-difluoro-2'-deoxycytidine 5'-triphosphate (dFdCyd/dFdCTP) would be altered after dCK gene transfer and that this change would result in an enhanced cytotoxic effect. To test this hypothesis, we examined dFdCyd/dFdCTP uptake and clearance in HT-29 human colon carcinoma xenografts in nude mice by high-performance liquid chromatography (HPLC) and fluorine-19 magnetic resonance spectroscopy (F-19 MRS).


METHODS
HT-29 tumors were grown from cells infected with either the retroviral vector alone (LNPO-LacZ) or vector containing the dCK gene (LNPO-dCK). HPLC and F-19 MRS analyses were performed after a single 160 mg/kg i.p. injection of dFdCyd. Tumor response was determined in animals receiving a similar dosing schedule of dFdCyd.


RESULTS
HPLC experiments revealed an increased tumor accumulation of dFdCTP in xenografts overexpressing dCK compared with wild-type controls (P < or = 0.05). dFdCTP in the dCK-infected tumors was easily identified at 24 h postinjection. Conversely, no dFdCTP could be detected in the control xenografts 14 h postinjection. Subsequent F-19 MRS experiments confirmed an altered uptake, revealing a 2.5-fold greater accumulation of dFdCyd/dFdCTP in the dCK xenografts. Whereas a modest tumor growth delay was observed in the wild-type tumors receiving dFdCyd, dCK xenografts demonstrated a marked tumor growth delay following treatment (P < or = 0.05).


CONCLUSIONS
These data support the hypothesis that increased expression of dCK cDNA in HT-29 xenografts results in an enhanced dFdCTP accumulation and prolonged elimination kinetics, and ultimately a potentiated in vivo tumor response to dFdCyd. Related to these effects, changes in the overall tumor metabolism of dFdCyd/dFdCTP was detectable by noninvasive F-19 MRS. These data are relevant to future preclinical and clinical studies evaluating dCK gene transfer and dFdCyd therapy.
Blackstock
A W
AW
Department of Radiation Oncology, Wake Forest University School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27156, USA. ablackst@wfubmc.edu
Lightfoot
H
H
Case
L D
LD
Tepper
J E
JE
Mukherji
S K
SK
Mitchell
B S
BS
Swarts
S G
SG
Hess
S M
SM
D000818:Animals; D002455:Cell Division / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D003570:Cytidine Triphosphate / Q000031:analogs & derivatives* / Q000378:metabolism; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000493:pharmacokinetics*; D003842:Deoxycytidine Kinase / Q000235:genetics* / Q000378:metabolism; D005260:Female; D005461:Fluorine; D015971:Gene Expression Regulation, Enzymologic; D018014:Gene Transfer Techniques; D019073:HT29 Cells; D006801:Humans; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D051379:Mice; D008819:Mice, Nude; D009374:Neoplasms, Experimental / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism*; D023041:Xenograft Model Antitumor Assays; D000093542:Gemcitabine
D003841:Deoxycytidine; C054403:2',2'-difluorodeoxycytidine 5'-triphosphate; D005461:Fluorine; D003570:Cytidine Triphosphate; D003842:Deoxycytidine Kinase; D000093542:Gemcitabine