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Development and Characterization of a Human Colon Adenocarcinoma Xenograft Deficient in Thymidine Salvage¹

Laboratories of Developmental Therapeutics, Division of Biochemical and Clinical Pharmacology, St. Jude Children's Research Hospital, Memphis, Tennessee 38101

In order to determine the contribution of thymidine (dThd) salvage to intrinsic resistance to antimetabolites (5-fluoropyrimidines, antifolates) in the human colon adenocarcinoma xenograft, HxGC₃, a subline deficient in thymidine kinase has been developed. A cell line (GC₃/M) was established in continuous culture that demonstrated a karyotype identical to that of the stem line of HxGC₃ (46,X,-Y+12). After inoculation of GC₃/M cells into immune-deprived CBA/CaJ mice, the HxGC₃/M xenografts retained histological, histochemical, and growth characteristics of the HxGC₃ xenograft. To develop a line deficient in dThd salvage, GC₃/M cells were selected with BrdUrd (100 µg/ml). Three clones characterized were unable to proliferate in HAT medium, and were deficient (<10% control) in the cytosolic form of thymidine kinase. Activities of dThd phosphorylase and dTMP synthase were unchanged from parental GC₃/M cells. Of the three clones inoculated into mice, GC₃/M TK^- 100 C₃ was tumorigenic, the xenografts demonstrating histological and growth characteristics similar to HxGC₃. The in vivo activity of the cytosolic form of dThd kinase was 3.5% of that in HxGC₃ xenografts. Incorporation in vivo of [methyl-³H]dThd into acid insoluble material was 14% of that in HxGC₃ tumors. Autoradiographs prepared from these tumors demonstrated that incorporation of radiolabel into nuclei occurred only in stromal cells derived from the host. It is anticipated that HxGC₃/M TK^- 100 C₃ will be a line valuable for determining the role of dThd salvage in intrinsic resistance to 5-fluoropyrimidines or antifolates in human colon adenocarcinomas growing as xenografts and also the relevance of dTMP synthase as a target for antimetabolites in this histiotype.

¹ Supported by Award CA 32613 from the National Cancer Institute and by American Lebanese Syrian Associated Charities.

² To whom requests for reprints should be addressed, at St. Jude Children's Research Hospital, Laboratories for Developmental Therapeutics, 332 North Lauderdale, P.O. Box 318, Memphis, TN 38101.

Received 7/22/86. Revised 11/10/86. Revised 1/ 2/86. Accepted 1/ 6/87.

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