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Use of isogenic human cancer cells for high-throughput screening and drug discovery


Cell-based screening for novel tumor-specific drugs has been compromised by the lack of appropriate control cells. We describe a strategy for drug screening based on isogenic human cancer cell lines in which key tumorigenic genes have been deleted by targeted homologous recombination. As a test case, a yellow fluorescent protein (YFP) expression vector was introduced into the colon cancer cell line DLD-1, and a blue fluorescent protein (BFP) expression vector was introduced into an isogenic derivative in which the mutant K-Ras allele had been deleted. Co-culture of both cell lines allowed facile screening for compounds with selective toxicity toward the mutant Ras genotype. Among 30,000 compounds screened, a novel cytidine nucleoside analog was identified that displayed selective activity in vitro and inhibited tumor xenografts containing mutant Ras. The present data demonstrate a broadly applicable approach for mining therapeutic agents targeted to the specific genetic alterations responsible for cancer development.

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Figure 1: Co-culture screening.
Figure 2: Structures of SC-D and TPT.
Figure 3: SC-D and TPT selectivity.
Figure 4: (A) TPT and (B) SC-D dose response.
Figure 5
Figure 6: Selectivity on K-Ras transformed rat kidney epithelial cells.
Figure 7: Effect of SC-D on tumor xenografts.


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We would like to thank the Developmental Therapeutics Program at the National Cancer Institute (NCI) for providing drug libraries and Dr. S. Shirasawa for providing the K-Ras KO cells. This work was supported by the Clayton Fund, the Miracle Foundation, the National Foundation for Cancer Research, and National Institutes of Health grants CA43460, CA09243, and CA62924.

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Correspondence to Bert Vogelstein.

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Torrance, C., Agrawal, V., Vogelstein, B. et al. Use of isogenic human cancer cells for high-throughput screening and drug discovery. Nat Biotechnol 19, 940–945 (2001).

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