Rapid Acquisition of Multicellular Drug Resistance After a Single Exposure of Mammary Tumor Cells to Antitumor Alkylating Agents

• Published in: JNCI : Journal of the National Cancer Institute 1994-07, Vol.86 (13), p.975-982
• Main Authors: Graham, Charles H., Kobayashi, Hiroaki, Stankiewicz, Krystyne S., Man, Shan, Kapitain, Sean J., Kerbel, R. S.
• Format: Article
• Language: English
• Subjects: Animals; Antineoplastic agents; Biological and medical sciences; Breast cancer; Breast Neoplasms - drug therapy; Breast Neoplasms - physiopathology; Chemotherapy; Cisplatin - pharmacology; Cyclophosphamide - analogs & derivatives; Cyclophosphamide - pharmacology; Drug Resistance; General aspects; Humans; Mammary Neoplasms, Experimental - drug therapy; Mammary Neoplasms, Experimental - physiopathology; Medical research; Medical sciences; Mice; Mice, Inbred BALB C; Pharmacology. Drug treatments; Time Factors; Tumor Cells, Cultured
• ISSN: 0027-8874; 1460-2105
• DOI: 10.1093/jnci/86.13.975

Background Clinical drug resistance is either intrinsic (de novo) or often acquired rapidly in conjunction with chemotherapy. By contrast, the selection of drug-resistant mutant cell lines in monolayer culture systems is usually a more protracted process. Sublines of mouse EMT-6 mammary tumor cells selected for resistance to various alkylating agents in vivo after serial passage into syngeneic mice manifest their resistance in vitro only when cultured as three-dimensional multicellular aggregates or spheroids. Purpose We examined whether a single exposure of mouse EMT-6 or human MDA-MB-231 breast cancer cells to alkylating agents in vitro is sufficient for the induction of a resistance phenotype, which may be detected by re-applying the drugs to cells grown as three-dimensional aggregates. Methods Mouse EMT-6 and human MDA-MB-231 breast cancer cells cultured as three-dimensional aggregates were exposed to a single dose of alkylating agent for 1–5 days. Aggregates were dispersed, and cells were plated as monolayer cultures for up to 8 weeks to allow for recovery. Colony-forming ability was assessed after a subsequent alkylating-agent exposure of cells cultured as monolayers or three-dimensional aggregates. Results: A single in vitro exposure to 12.5-μM cisplatin (CDDP) for 5 days or 25 μM 4-hydroperoxycyclophosphamide (4-O2H CTX) for 1 or 3 days without changing the medium was sufficient to induce transient but substantial resistance in EMT-6 cells as determined by clonogenic assays. Such resistance was not detected when monolayer cell cultures were used. The concentration of 4-O2H-CTX and the length of time the cells remained in three-dimensional culture after initial exposure to this drug was associated with the degree of subsequent drug resistance of cells grown as three-dimensional cultures. Furthermore, this acquired resistance after a single drug exposure was accompanied by changes in the three-dimensional architecture of the cell aggregates, which now formed much more compact multicellular spheroids. Similarly, a single exposure to 4-O2H-CTX was enough to bring about resistance in MDA-MB-231 cells detectable only in three-dimensional cultures, as well as the change in three-dimensional architecture. Conclusions Rapid acquisition of resistance likely represents a physiologic mechanism of adaptation operative at the multicellular level rather than a stable genetic change and may be one of the reasons for the rapid development of drug resistance acquire