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.
Torrance
C J
CJ
The Howard Hughes Medical Institute, 1650 Orleans Street, Baltimore, MD 21231, USA.
Agrawal
V
V
Vogelstein
B
B
Kinzler
K W
KW
D000736:Androstenes / Q000494:pharmacology; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D001426:Bacterial Proteins; D004354:Drug Screening Assays, Antitumor / Q000379:methods*; D005260:Female; D017353:Gene Deletion; D011905:Genes, ras; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins; D051379:Mice; D018345:Mice, Knockout; D008819:Mice, Nude; D016296:Mutagenesis; D008926:Plicamycin / Q000494:pharmacology; D011995:Recombination, Genetic; D013457:Sulfur Compounds / Q000494:pharmacology; D013778:Tetrazolium Salts / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D000736:Androstenes; D000970:Antineoplastic Agents; D001426:Bacterial Proteins; D008164:Luminescent Proteins; D013457:Sulfur Compounds; D013778:Tetrazolium Salts; C409504:blue fluorescent protein, Aequorea victoria; C054562:yellow fluorescent protein, Bacteria; D049452:Green Fluorescent Proteins; C070594:demethoxyviridin; C009591:triphenyltetrazolium; D008926:Plicamycin