The "comparative growth assay": examining the interplay of anti-cancer agents with cells carrying single gene alterations.
We have developed a "comparative growth assay" that complements current assays of drug effects based on cytotoxicity. A co-culture of two cell lines, one of which is fluorescently labeled, is exposed to a cytotoxic agent and the proportion of fluorescent cells is compared with that of a baseline unexposed co-culture. For demonstration purposes, two HCT116 cell lines (an hMLH1 homozygous and an hMLH1 heterozygous mutant), altered by insertion of vector alone or the same vector carrying an insert for the expression of enhanced green fluorescent protein (EGFP), were exposed to numerous "anti-cancer" agents. The assay was further validated in a system of two cell lines differing only in the expression of the breast cancer resistance protein (BRCP). The assay allowed the estimation of the duration of action of a particular agent. Assessment of the agent's differential activity over a given time in culture could be expressed as a selection rate, which we chose to describe on an "average selection per day" basis. We conclude that this assay: 1) provides insight into the differential dynamic effects of chemotherapeutic agents or radiation; and 2) allows, through the use of matched cell lines, the investigation of critical physiologic features that govern cell sensitivity.
Marlene and Stewart Greenebaum Cancer Center, University of Maryland Medicine, Baltimore 21201-1595, USA.
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Hausner
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Venzon
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Grogan
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Kirsch
D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000096:biosynthesis / Q000235:genetics; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000494:pharmacology; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods*; D002455:Cell Division / Q000187:drug effects / Q000528:radiation effects; D018920:Coculture Techniques; D003110:Colonic Neoplasms / Q000235:genetics / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004307:Dose-Response Relationship, Radiation; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D005434:Flow Cytometry; D005456:Fluorescent Dyes / Q000494:pharmacology; D005822:Genetic Vectors; D005839:Gentamicins / Q000494:pharmacology; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000378:metabolism; D008769:Methylnitronitrosoguanidine / Q000494:pharmacology; D008942:Mitoxantrone / Q000494:pharmacology; D009154:Mutation; D009363:Neoplasm Proteins; D013997:Time Factors; D014162:Transfection; D014407:Tumor Cells, Cultured
C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D000900:Anti-Bacterial Agents; D000970:Antineoplastic Agents; D005456:Fluorescent Dyes; D005839:Gentamicins; D008164:Luminescent Proteins; D009363:Neoplasm Proteins; D008769:Methylnitronitrosoguanidine; D049452:Green Fluorescent Proteins; C010680:antibiotic G 418; D008942:Mitoxantrone